US20160068863A1

US20160068863A1 - Methods for heart regeneration

Info

Publication number: US20160068863A1
Application number: US14/946,644
Authority: US
Inventors: Aitor Aguirre; Ignacio Sancho-Martinez; Juan Carlos Izpisua-Belmonte
Original assignee: Salk Institute for Biological Studies
Current assignee: Salk Institute for Biological Studies
Priority date: 2012-10-11
Filing date: 2015-11-19
Publication date: 2016-03-10
Also published as: US20140221463A1; US9220721B2

Abstract

Methods for heart regeneration are provided. The invention provided herein includes methods of modulating proliferation of cardiomyocytes using small molecules and micro RNAs. In embodiments, the methods provided may be used to increase proliferation or cardiomyocytes. Further provided are methods to be used for the treatment of myocardial infarction.

Description

CROSS-REFERENCES TO RELATED APPLICATIONS

This is a divisional of U.S. patent application Ser. No. 14/052,538, filed Oct. 11, 2013, which claims the benefit of U.S. Provisional Appl. No. 61/712,701, filed Oct. 11, 2012. The prior applications are incorporated herein in their entirety.

BACKGROUND OF THE INVENTION

Heart failure remains one of the leading causes of mortality in the developed world. Whereas the mammalian heart is endowed with certain regenerative potential, endogenous cardiomyocyte proliferation is insufficient for functional heart repair upon injury. Interestingly, non-mammalian vertebrates, such as the zebrafish, can regenerate the damaged heart by inducing cardiomyocyte dedifferentiation and proliferation. By screening regenerating zebrafish hearts Applicants identified miR-99/100 down-regulation as a key process driving cardiomyocyte dedifferentiation. Experimental down-regulation of miR-99/100 in primary adult murine and human cardiomyocytes led to an increase in the number of proliferating cardiomyocytes. AAV-mediated in vivo down-regulation of miR-99/100 after acute myocardial injury in mice induced mature cardiomyocyte proliferation, diminished infarct size and improved heart function. Applicants' study unveils conserved regenerative mechanisms between zebrafish and mammalian cardiomyocytes and represents a proof-of-concept on the suitability of activating pro-regenerative responses for healing the diseased mammalian heart.

BRIEF SUMMARY OF THE INVENTION

In one aspect, a method of modulating proliferation of a cardiomyocyte is provided. The method includes (i) transfecting a cardiomyocyte with a nucleic acid encoding a micro RNA modulator, thereby forming a transfected cardiomyocyte; and (ii) allowing the transfected cardiomyocyte to divide, thereby modulating proliferation of the cardiomyocyte.
In another aspect, a method of modulating proliferation of a cardiomyocyte is provided. The method includes (i) contacting a cardiomyocyte with a small molecule, thereby forming a treated cardiomyocyte; and (ii) allowing the treated cardiomyocyte to divide, thereby modulating proliferation of the cardiomyocyte.
In another aspect, a method of treating myocardial infarction in a subject in need thereof is provided. The method includes administering to the subject a therapeutically effective amount of a nucleic acid encoding a micro RNA modulator, wherein the RNA modulator increases cardiomyocyte proliferation thereby treating the myocardial infarction.
In another aspect, a method of treating myocardial infarction in a subject in need thereof is provided. The method includes administering to the subject a therapeutically effective amount of a nucleic acid encoding an antagonist of a mir 99 micro RNA and a nucleic acid encoding an antagonist of a let-7a micro RNA, thereby treating the myocardial infarction.
In another aspect, a method of treating myocardial infarction in a subject in need thereof is provided. The method includes administering to the subject a therapeutically effective amount of a small molecule, wherein the small molecule increases cardiomyocyte proliferation thereby treating the myocardial infarction.

BRIEF DESCRIPTION OF THE DRAWINGS

FIGS. 1A-1J. miR-99/100 and Let-7a/c are involved in the early cardiac regenerative response in the zebrafish. (FIG. 1A) Amputated hearts were allowed to regenerate for 3 and 7 days, and then analyzed by real time RT-PCR for microRNA candidates miR-99/100 and Let-7a/c (n=8). (FIGS. 1B, C) FISH/double immunofluorescence was used to determine cardiomyocyte specific expression of microRNA-100 (n=5, see also FIG. 6) and its downstream targets fntβ (FIG. 1B) and smarca5 (FIG. 1C) in uninjured (left panel) and 7 dpa conditions (right panel). Cardiomyocytes in regenerating hearts exhibited remarkable low levels of both miRs, and inversely correlating high levels of Fntβ and Smarca5. (FIG. 1D) Gene expression fold change of fntβ and smarca5 in amputated hearts (n=8). (FIG. 1E) Chemical inhibition of fnt activity with Tipifarnib dramatically reduced cardiomyocyte proliferation and heart regeneration in amputated fish, leading to scarring as determined by Masson's Trichromic (FIG. 1F) staining and BrDU incorporation (FIG. 1G) (n=6). (FIGS. 1H, I, J) Knock-down of fntβ and/or smarca5 in embryos resulted in abnormally small animals and reduced ventricle size in cm1c2:GFP animals. The same phenotype was observed upon injection of miR-99/100 mimics (n>50). Dashed line: amputation plane. Boxed area: magnified field. Arrowheads: cells of interest.

FIGS. 2A-2J. Heart regeneration in the fish is controlled by miR-99/100 and Let-7a/c. (FIG. 2A,B) Dedifferentiating cardiomyocytes express high levels of Fntβ (from top to bottom, uninjured, 3 dpa and 7 dpa, panel FIG. 2A) and Smarca5 (from top to bottom, uninjured, 3 dpa and 7 dpa, panel FIG. 2B) with a 3.5-fold and 7-fold protein increment at 7 dpa, respectively, (see histogram in FIGS. 2C, D; n=5). (FIGS. 2E, F, G) Exogenous intra-cardiac administration of miR-99/100 mimics led to defective cardiac regeneration, scarring and reduced cardiomyocyte proliferation (n=6). (FIG. 2H) miR-99/100 inhibitors exerted the opposite effect in uninjured adult animals, inducing significant cardiac hyperplasia (FIG. 2I) and increased ventricle size (FIG. 2J; n=6). Dashed line: amputation plane. Boxed area: magnified area. Ec: endocardial cells; cm: cardiomyocytes. Arrowheads: cells of interest.

FIGS. 3A-3H. Adult mammalian cardiomyocytes can be directed to a proliferative state by forced silencing of the miR-99/100 cluster. Expression patterns of the miR-99/100 cluster during development in murine (FIG. 3A) and human cardiomyocytes (FIG. 3C) were determined by qRT-PCR (n=6). (FIGS. 3B, D) Dynamics of cardiomyocyte progenitor marker and Fntβ/Smarca5 expression during murine (FIG. 3B) and human (FIG. 3D) cardiac maturation. Both Fntβ and Smarca5 are up-regulated at cardiac progenitor stages, and strongly repressed in adult hearts (n=6). (FIGS. 3E, F) Cultured adult murine cardiomyocytes transduced with anti-miR-99/100+anti-Let-7 reverted to a dedifferentiated-like state, re-expressing GATA4 and dissembling the cytoskeleton (see also FIG. 17; n=3). (FIGS. 3G, H) Transduction with anti-miRs was sufficient to efficiently drive adult cardiomyocytes to a proliferative state (FIG. 3G), with beating functionality (FIG. 3H) (n=3).

FIGS. 4A-4L. MicroRNA silencing is sufficient to induce heart regeneration in a murine model of myocardial infarction. (FIGS. 4A, B) Organotypic cultures of adult heart tissue were employed to study the effects of microRNA-99/100 and Let-7a/c silencing. (FIG. 4A) After 7 days of treatment, myocardial tissue became disorganized, with a loss of sarcomeric structures as determined by electron microscopy analysis (n=6). (FIG. 4B) Furthermore, re-expression of dedifferentiation markers was detected by immunofluorescence evaluation/quantification, as well as cardiomyocyte proliferation and FNTβ and SMARCA5 expression in normoxic and hypoxic conditions (n=6). (FIG. 4C) Representative pictures of echocardiography performed in control (left panel) and treated animals (right panel) at 18 days post-myocardial infarction (MI). In vivo silencing of miR-99/100 and Let-7 led to significant improvements in fractional shortening (FIGS. 4D, F) and ejection fraction (FIGS. 4E, F) at 18 days post-MI (n=5). (FIG. 4G) Left panel, reduced infarct size in miR-99/100 and Let-7 treated animals is confirmed by Masson's trichromic staining; (FIG. 4G) Right panel, quantification of scar size by Masson's thrichromic staining (FIG. 4G; n=5). (FIGS. 4H-J) Recovery was accompanied by cardiomyocyte-specific expression of FNTβ (FIG. 4H) and SMARCA5 (FIG. 4I), as well as GATA4 re-expression (FIG. 4J, right panel shows quantification of this data) as determined by immunofluorescence analyses (n=5). (FIGS. 4K, L, panel on the right shows quantification of this data) Regeneration was mediated by a dramatic increase in proliferating cardiomyocytes as determined by PCNA (FIG. 4K) and H3P (FIG. 4L) stainings (n=5). Arrowheads: cells of interest.

FIGS. 5A-5C. miR-99/100 and Let-7a/c are located in same genomic cluster and their functions and protein targets are conserved across vertebrates. (FIG. 5A) Microarray analysis identified a subset of differentially regulated microRNAs during early stages of regeneration in the zebrafish heart (n=4). Interestingly, most of them were consistently down-regulated. (FIG. 5B) Bioinformatic analysis of the most relevant signaling pathways targeted by miR-99/100 and Let-7a/c. (FIG. 5C) Genomic organization and conservation of miR-99/100 and Let-7a/c clusters in vertebrates (upper left: zebrafish miR-100 cluster; upper right: zebrafish miR-99 cluster; lower left: human miR-99/100 clusters; lower right: murine miR-99/100 clusters).

FIGS. 6A-6D. Expression of miR-99/100 is restricted to cardiomyocytes and inversely correlates with Fntβ and Smarca5. (FIGS. 6A-D) FISH/immunofluorescence analyses on zebrafish heart sections from uninjured and amputated animals at 3 and/or 7 dpa. (FIGS. 6A, B) FISH/immunofluorescence analysis of miR-100 and Fntβ/Smarca5 expression (see also FIG. 1; n=8). (FIGS. 6C, D) FISH/immunofluorescent analysis of miR-99 and Fntβ/Smarca5 expression during regeneration (upper row, miR-99 effects on Fntb expression at uninjured (left), 3 dpa (center) and 7 dpa (right) conditions; lower row, miR-99 effects on Smarca5 expression at uninjured (left), 3 dpa (center) and 7 dpa (right) conditions). (n=8). Dashed line: amputation plane. Boxed area: magnified in inset.

FIGS. 7A-7B. Relevant protein targets of interest in heart regeneration regulated by miR-99/100 and Let-7a/c. (FIG. 7A) Miranda-based binding predictions of miR-99/100 to zebrafish Fntb (upper panel) and Smarca5 (lower panel) 3′ UTRs. (FIG. 7B) Luciferase assay to determine biochemical binding of miR-99/100 to the predicted targets Fntb and Smarca5 for zebrafish (upper and middle rows) and human (lower row) 3′UTRs. Fish and human UTRs were subcloned in the pGL3 vector and subjected to microRNA mimic knockdown in vitro. pGL3: empty vector; AS-UTR: antisense-UTR (negative control); UTR: 3′untranslated region.

FIGS. 8A-8B. Fntα, the structural subunit of Fnt, was constitutively expressed in cardiomyocytes regardless of regeneration conditions. Fnta expression was determined by immunofluorescence (FIG. 8A) and qRT-PCR (FIG. 8B, n=4). Dashed line: amputation plane. Boxed area: magnified section.

FIGS. 9A-9D. MiR-99/100 plays a role in heart development. (FIG. 9A) Expression of miR-99/100 is very low during the first stages of development and dramatically increases at 3 dpf in zebrafish (n=10). (FIG. 9B) fntβ and smarca5 expression inversely correlate with miR-99/100 in developing embryos (n=10). (FIGS. 9C, D) Both Fntβ and Smarca5 are present at high levels in developing hearts (n=10). V: ventricle; A: atrium; Eb: erythroblasts.

FIGS. 10A-10D. Targets of miR-99/100 and Let-7a/c return to basal levels of expression after cardiomyocytes come back to their quiescent state. (FIGS. 10A, B) Immunofluorescent stainings for proliferating cardiomyocytes at 30 dpa, when regeneration in the zebrafish heart is mostly complete (n=3). Fntβ and Smarca5 presence in the myocardium returned to pre-amputation levels at 30 dpa, when regeneration is mostly completed. (FIG. 10C) Strategy for the scarring experiments to identify the importance in regeneration of miR-99/100. (FIG. 10D) Successful heart delivery of antagomiRs was evaluated by injection of a matched-size Cy5-labelled oligonucleotide against GFP in cm1c2:GFP animals (n=3).Dashed line: amputation plane. Boxed area: magnified section.

FIGS. 11A-11B. Direct substrates of fnt are activated in regenerating hearts. (FIGS. 11A, B) At 3 and 7 dpa, a significant fraction of farnesylated proteins was detected in dedifferentiating cardiomyocytes as determined by immunofluorecence (n=5). Boxed area: magnified section.

FIGS. 12A-12F. Signaling downstream of fntβ in the MAPK signaling pathway were consistently up-regulated in regenerating hearts. Ras (FIGS. 12A, B, C) and c-myc (FIGS. 12D, E, F), activators of the MAPK pathway regulated by miR-99/100 and Let-7a/c were up-regulated as determined by immunofluorescence (FIGS. 12A,D) and by qRT-PCR (FIGS. 12B,C,E,F; n=5). Dashed line: amputation plane. Boxed area: magnified section. Arrowheads: cells of interest.

FIGS. 13A-13B. Chromatin remodeling is a necessary step in cardiomyocyte dedifferentiation. (FIGS. 13A,B) The chromatin remodeling agents Cbx5 (FIG. 13A) and Cbx3a (FIG. 13B), which act in concert with Smarca5, were found in the nucleus of dedifferentiating cardiomyocytes indicating a wave of chromatin remodeling. The following is shown in the histograms of FIG. 13A and FIG. 13B from left to right: First panel (bottom and top histogram): DAPI stain, second panel (bottom and top histogram): MyHC stain; third panel (bottom and top histogram): PCNA stain; forth panel (bottom and top histogram): Cbx5 stain; fifth panel (bottom and top histogram): merge of histograms of panels one, two three and four. (n=4). Arrowheads: cells of interest.

FIG. 14. Expression of miR-99/100 cluster is a switch to promote or inhibit cardiomyocyte dedifferentiation and proliferation in vertebrates. In the proposed model of action, quiescent cardiomyocytes express high levels of miR-99/100 and Let-7a/c, which inhibit the expression of key members of the proliferative activation cascade (Uninjured condition shown in left panel). Upon injury, cardiomyocytes cease to express these miRs, leading to the over-expression of key regulators of two parallel pathways simultaneously: MAPK signaling and chromatin remodeling (Injury condition shown in right panel). These changes lead to a proliferation-permissive state in cardiomyocytes, which become more receptive towards proliferative signals coming from the injured area and proliferate to replenish the lost tissue. The symbols “X” indicate blocked pathways.

FIGS. 15A-15D. FISH/IF for different developmental stages in the mouse heart. (FIGS. 15A-C) Identical patterns of expression were found for miR-99/100 and Let-7a/c, as well as Fntβ and Smarca5, to those observed in the developing zebrafish (FIG. 15A: embryonic day 11; FIG. 15B: postnatal day 2; FIG. 15C: adult) (n=6). (FIG. 15D) Quantification showing the number of double positive cells for miR-99/100 (FIG. 15D upper left), Let-7a/c (FIG. 15D upper right), Fntβ (FIG. 15D lower left) and Smarca5 (FIG. 15D lower right) (n=6).

FIGS. 16A-16B. Human ESC-derived, proliferation-competent immature cardiomyocytes (hiCM) possessed the same phenotype observed in fish and mouse dedifferentiated cardiomyocytes. (FIG. 16A) Immunofluorescence for the indicated proteins in human embryonic stem cells (FIG. 16A, Images are individual panels of the merged figure shown at in the last column). (FIG. 16B) hiCMs showed expression of GATA4, SMARCA5 and FNTβ, which was progressively lost with decreased proliferative capacity (n=3). The following is shown in the histograms of FIG. 16A and FIG. 16B from left to right: Histograms show immunostaining with DAPI (first panel), MyHC (second panel), GATA4 (third panel), SMARCA5 (forth panel top), FNTbeta (forth panel middle), of H3P (forth panel bottom) and a merged histogram (fifth panel).

FIGS. 17A-17F. miR silencing leads to dedifferentiation and proliferation of cardiomyocytes. (FIG. 17A, Images represent individual panels of the merged image shown in the last column) Untreated adult murine cardiomyocytes spontaneously disorganized sarcomeric structures in vitro, but did not dedifferentiate or express FNTβ, SMARCA5, GATA4 or proliferative markers. However, upon lentiviral transduction with anti-Let-7 alone (FIG. 17B, Images represent individual panels of the merged image shown in the last column) or both anti-Let-7 and anti-miR-99/100 (FIG. 17C, Images represent individual panels of the merged image shown in the last column) they re-expressed all those markers (n=3). (FIG. 17D) Functional beating properties were preserved in dedifferentiated cardiomyocytes, suggesting a degree of spontaneous redifferentiation, except for anti-Let-7 treatment. (FIGS. 17E, F) Cardiomyocyte dedifferentiation was evaluated by simultaneously measuring GATA4 and sarcomeric myosin expression and organization in cultured cardiomyocytes (n=3).

FIGS. 18A-18C. microRNA silencing is specific for cardiomyocytes. (FIG. 18A) Human Fibroblasts (left panel) or endothelial cells (right panel) expressed basal levels of FNTβ, SMARCA5 and negligible levels of miR-99/100 and Let-7a/c (data not shown), and were insensitive to miR silencing (FIGS. 18A, B, C), indicating specificity of the treatment in a heart setting. (FIG. 18C) Images used for quantification of data shown in A and B (Images represent individual panels of the merged image shown in the last column) (n=3).

FIGS. 19A-19B (from top to bottom, confocal analysis of FNTB, SMARCA5, Cx43, GATA4 and H3P). Histograms to the left represent positional information from the numbers shown in the insets. Ex vivo organotypic culture of murine myocardial tissue reveals sustained cardiomyocyte proliferation. Adult mouse heart tissue was cultured and treated with empty vector (pmiRZip), anti-miR-99/100 or both anti-miR-99/100 and anti-Let-7 (lentiviral activation was followed with a GFP reporter). Confocal analysis after 7 days of miR silencing led to significantly increased FNTβ and SMARCA5 (FIGS. 19A, B), enhanced numbers of dedifferentiated cardiomyocytes —determined by Cx43 and GATA4 expression— (FIG. 19B) and significantly increased number of proliferating cells (n=4). Dashed line and corresponding numbers: nuclear profile of representative cells. Boxed area: magnified section.

FIGS. 20A-20E. Hypoxic injury in organotypic culture leads to increased dedifferentiation. (FIG. 20A) Schematic of hypoxia experiments. (FIG. 20B) Efficient lentiviral delivery was achieved in ex vivo conditions for all anti-miRs. (FIGS. 20C, D) Histomorphometric evaluation of the damaged myocardium in normoxic and hypoxic conditions by employing Masson's trichrome staining (n=4). (FIG. 20E) The dedifferentiation response was characterized by GATA4, Cx43, H3P, Fntβ and Smarca5 stainings (From top to bottom, Cx43, SMARCA5, H3P) (n=4). Dashed line: adjacent necrotic patch. Boxed area: magnified section. Dashed lines: necrotic tissue.

FIGS. 21A-21B. Echocardiography showing functional improvement in infarcted mice treated with anti-miR-99/100 and anti-Let-7a (upper panel) and quantification of that data (lower panel). Animals were subjected to LAD artery ligation to provike infarction followed by tintramyocardial administration of antimiR containing AAV2/9 vectors. Functional heart recovery was measured versus untreated contrail animals. (FIG. 21A) At 18 dpi (days post infarction) mice treated with antimiR's exhibit a significant improvement in ejection fraction and fractional shortening, as well as reduction in scar size. (FIG. 21B) At 3 mpi (months post infarction) the improvements are still present (left panels, echocardiographic quantification; right panel, representative images), with further reduction of scar size and significant enlargement of the LAVW, indicative of replenishment of the myocardial mass.

FIGS. 22A-22B. In vivo regenerative reprogramming by anti-miR delivery (FIG. 22A). In vivo-induced cardiomyocyte proliferation by anti-miR delivery (18 days post-infarction) (left panels, PCNA and H3P staining showing proliferating cardiomyocytes; right panel, quantification of the data) (FIG. 22B).

FIGS. 23A-23C. In vivo dedifferentiation by anti-miR delivery. (FIG. 23A): left panel, GATA4 immunofluorescence; right panel, quantification of the data shown (FIG. 23B): histogram showing percentage of dedifferentiated CMs; (FIG. 23C): SMARCA5 immunofluorescence.

FIGS. 24A-24B. Anti-miR99/100/let-7 delivery enhances functional recovery after infarction. Echocardiography measurements in infarcted animals treated with control vs. anti-miR treatment indicate significant regeneration after treatment (FIG. 24A). Qunatification of those animals shows statistically significant functional recovery, both in fraction shortening (left panel) as well as ejection fraction (right panel).

FIGS. 25A-25B. Anti-miR99/100/let-7 delivery enhances functional recovery after infarction. Heart function improvement by anti-miR treatment is sustained over long periods of time (ejection fraction) and seems to involve regeneration of the myocardial mass (LAWV thickening, immunohistological analysis shown in FIG. 25A). Ultrasound analysis shown in FIG. 25B (abbreviations AW; anterior wall; ID: internal diameter; reflects heart dilation; PW: posterior wall).

REFERENCE TO A “SEQUENCE LISTING,” A TABLE, OR A COMPUTER PROGRAM LISTING APPENDIX SUBMITTED AS AN ASCII TEXT FILE

The Sequence Listing written in file 7158-94960-03_Sequence_Listing, created on Nov. 18, 2015, 7.45 MB, machine format IBM-PC, MS Windows operating system, is hereby incorporated by reference.

DETAILED DESCRIPTION OF THE INVENTION

Definitions

While various embodiments and aspects of the present invention are shown and described herein, it will be obvious to those skilled in the art that such embodiments and aspects are provided by way of example only. Numerous variations, changes, and substitutions will now occur to those skilled in the art without departing from the invention. It should be understood that various alternatives to the embodiments of the invention described herein may be employed in practicing the invention.
The section headings used herein are for organizational purposes only and are not to be construed as limiting the subject matter described. All documents, or portions of documents, cited in the application including, without limitation, patents, patent applications, articles, books, manuals, and treatises are hereby expressly incorporated by reference in their entirety for any purpose.
Unless defined otherwise, technical and scientific terms used herein have the same meaning as commonly understood by a person of ordinary skill in the art. See, e.g., Singleton et al., DICTIONARY OF MICROBIOLOGY AND MOLECULAR BIOLOGY 2nd ed., J. Wiley & Sons (New York, N.Y. 1994); Sambrook et al., MOLECULAR CLONING, A LABORATORY MANUAL, Cold Springs Harbor Press (Cold Springs Harbor, N.Y. 1989). Any methods, devices and materials similar or equivalent to those described herein can be used in the practice of this invention. The following definitions are provided to facilitate understanding of certain terms used frequently herein and are not meant to limit the scope of the present disclosure.
“Nucleic acid” refers to deoxyribonucleotides or ribonucleotides and polymers thereof in either single- or double-stranded form, and complements thereof. The term “polynucleotide” refers to a linear sequence of nucleotides. The term “nucleotide” typically refers to a single unit of a polynucleotide, i.e., a monomer. Nucleotides can be ribonucleotides, deoxyribonucleotides, or modified versions thereof. Examples of polynucleotides contemplated herein include single and double stranded DNA, single and double stranded RNA (including siRNA), and hybrid molecules having mixtures of single and double stranded DNA and RNA. The terms also encompass nucleic acids containing known nucleotide analogs or modified backbone residues or linkages, which are synthetic, naturally occurring, and non-naturally occurring, which have similar binding properties as the reference nucleic acid, and which are metabolized in a manner similar to the reference nucleotides. Examples of such analogs include, without limitation, phosphorothioates, phosphoramidates, methyl phosphonates, chiral-methyl phosphonates, and 2-O-methyl ribonucleotides.
A “miRNA” or “microRNA” as provided herein refers to a nucleic acid that forms a double stranded RNA, which double stranded RNA has the ability to reduce or inhibit expression of a gene or target gene when expressed in the same cell as the gene or target gene. The complementary portions of the nucleic acid that hybridize to form the double stranded molecule typically have substantial or complete identity. In one embodiment, a microRNA refers to a nucleic acid that has substantial or complete identity to a target gene and forms a double stranded miRNA. In embodiments, the miRNA inhibits gene expression by interacting with a complementary cellular mRNA thereby interfering with the expression of the complementary mRNA. Typically, the nucleic acid is at least about 15-50 nucleotides in length (e.g., each complementary sequence of the double stranded miRNA is 15-50 nucleotides in length, and the double stranded miRNA is about 15-50 base pairs in length). In other embodiments, the length is 20-30 base nucleotides, preferably about 20-25 or about 24-29 nucleotides in length, e.g., 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, or 30 nucleotides in length.
The words “complementary” or “complementarity” refer to the ability of a nucleic acid in a polynucleotide to form a base pair with another nucleic acid in a second polynucleotide. For example, the sequence A-G-T is complementary to the sequence T-C-A. Complementarity may be partial, in which only some of the nucleic acids match according to base pairing, or complete, where all the nucleic acids match according to base pairing.
The terms “protein”, “peptide”, and “polypeptide” are used interchangeably to denote an amino acid polymer or a set of two or more interacting or bound amino acid polymers. The terms apply to amino acid polymers in which one or more amino acid residue is an artificial chemical mimetic of a corresponding naturally occurring amino acid, as well as to naturally occurring amino acid polymers and non-naturally occurring amino acid polymer.
The term “amino acid” refers to naturally occurring and synthetic amino acids, as well as amino acid analogs and amino acid mimetics that function in a manner similar to the naturally occurring amino acids. Naturally occurring amino acids are those encoded by the genetic code, as well as those amino acids that are later modified, e.g., hydroxyproline, γ-carboxyglutamate, and O-phosphoserine. Amino acid analogs refers to compounds that have the same basic chemical structure as a naturally occurring amino acid, i.e., an a carbon that is bound to a hydrogen, a carboxyl group, an amino group, and an R group, e.g., homoserine, norleucine, methionine sulfoxide, methionine methyl sulfonium. Such analogs have modified R groups (e.g., norleucine) or modified peptide backbones, but retain the same basic chemical structure as a naturally occurring amino acid. Amino acid mimetics refers to chemical compounds that have a structure that is different from the general chemical structure of an amino acid, but that functions in a manner similar to a naturally occurring amino acid. The terms “non-naturally occurring amino acid” and “unnatural amino acid” refer to amino acid analogs, synthetic amino acids, and amino acid mimetics which are not found in nature.
Amino acids may be referred to herein by either their commonly known three letter symbols or by the one-letter symbols recommended by the IUPAC-IUB Biochemical Nomenclature Commission. Nucleotides, likewise, may be referred to by their commonly accepted single-letter codes.
“Conservatively modified variants” applies to both amino acid and nucleic acid sequences. With respect to particular nucleic acid sequences, conservatively modified variants refers to those nucleic acids which encode identical or essentially identical amino acid sequences, or where the nucleic acid does not encode an amino acid sequence, to essentially identical sequences. Because of the degeneracy of the genetic code, a large number of functionally identical nucleic acids encode any given protein. For instance, the codons GCA, GCC, GCG and GCU all encode the amino acid alanine. Thus, at every position where an alanine is specified by a codon, the codon can be altered to any of the corresponding codons described without altering the encoded polypeptide. Such nucleic acid variations are “silent variations,” which are one species of conservatively modified variations. Every nucleic acid sequence herein which encodes a polypeptide also describes every possible silent variation of the nucleic acid. One of skill will recognize that each codon in a nucleic acid (except AUG, which is ordinarily the only codon for methionine, and TGG, which is ordinarily the only codon for tryptophan) can be modified to yield a functionally identical molecule. Accordingly, each silent variation of a nucleic acid which encodes a polypeptide is implicit in each described sequence with respect to the expression product, but not with respect to actual probe sequences.
The terms “identical” or percent “identity,” in the context of two or more nucleic acids or proteins, refer to two or more sequences or subsequences that are the same or have a specified percentage of nucleotides or amino acids that are the same (i.e., about 60% identity, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or higher identity over a specified region, when compared and aligned for maximum correspondence over a comparison window or designated region) as measured using a BLAST or BLAST 2.0 sequence comparison algorithms with default parameters described below, or by manual alignment and visual inspection. See e.g., the NCBI web site at ncbi.nlm.nih.gov/BLAST. Such sequences are then said to be “substantially identical.” This definition also refers to, or may be applied to, the compliment of a test sequence. The definition also includes sequences that have deletions and/or additions, as well as those that have substitutions. Identity typically exists over a region that is at least about 50 amino acids or nucleotides in length, or over a region that is 50-100 amino acids or nucleotides in length, or over the entire length of a given sequence.
The term “gene” means the segment of DNA involved in producing a protein; it includes regions preceding and following the coding region (leader and trailer) as well as intervening sequences (introns) between individual coding segments (exons). The leader, the trailer as well as the introns include regulatory elements that are necessary during the transcription and the translation of a gene. Further, a “protein gene product” is a protein expressed from a particular gene.
The word “expression” or “expressed” as used herein in reference to a gene means the transcriptional and/or translational product of that gene. The level of expression of a DNA molecule in a cell may be determined on the basis of either the amount of corresponding mRNA that is present within the cell or the amount of protein encoded by that DNA produced by the cell (Sambrook et al., 1989, Molecular Cloning: A Laboratory Manual, 18.1-18.88).
The term “recombinant” when used with reference, e.g., to a cell, or nucleic acid, protein, or vector, indicates that the cell, nucleic acid, protein or vector, has been modified by the introduction of a heterologous nucleic acid or protein or the alteration of a native nucleic acid or protein, or that the cell is derived from a cell so modified. Thus, for example, recombinant cells express genes that are not found within the native (non-recombinant) form of the cell or express native genes that are otherwise abnormally expressed, under expressed or not expressed at all.
The term “heterologous” when used with reference to portions of a nucleic acid or protein indicates that the nucleic acid or protein comprises two or more subsequences that are not found in the same relationship to each other in nature. For instance, the nucleic acid is typically recombinantly produced, having two or more sequences from unrelated genes arranged to make a new functional nucleic acid, e.g., a promoter from one source and a coding region from another source. Similarly, a heterologous protein indicates that the protein comprises two or more subsequences that are not found in the same relationship to each other in nature (e.g., a fusion protein).
The term “exogenous” refers to a molecule or substance (e.g., nucleic acid or protein) that originates from outside a given cell or organism. Conversely, the term “endogenous” refers to a molecule or substance that is native to, or originates within, a given cell or organism.
A “vector” is a nucleic acid that is capable of transporting another nucleic acid into a cell. A vector is capable of directing expression of a protein or proteins encoded by one or more genes carried by the vector when it is present in the appropriate environment.
A “viral vector” is a viral-derived nucleic acid that is capable of transporting another nucleic acid into a cell. A viral vector is capable of directing expression of a protein or proteins encoded by one or more genes carried by the vector when it is present in the appropriate environment. Examples for viral vectors include, but are not limited to retroviral, adenoviral, lentiviral and adeno-associated viral vectors.
A “cell culture” is an in vitro population of cells residing outside of an organism. The cell culture can be established from primary cells isolated from a cell bank or animal, or secondary cells that are derived from one of these sources and immortalized for long-term in vitro cultures.
The terms “culture,” “culturing,” “grow,” “growing,” “maintain,” “maintaining,” “expand,” “expanding,” etc., when referring to cell culture itself or the process of culturing, can be used interchangeably to mean that a cell is maintained outside the body (e.g., ex vivo) under conditions suitable for survival. Cultured cells are allowed to survive, and culturing can result in cell growth, differentiation, or division. The term does not imply that all cells in the culture survive or grow or divide, as some may naturally sense, etc. Cells are typically cultured in media, which can be changed during the course of the culture.
The terms “media” and “culture solution” refer to the cell culture milieu. Media is typically an isotonic solution, and can be liquid, gelatinous, or semi-solid, e.g., to provide a matrix for cell adhesion or support. Media, as used herein, can include the components for nutritional, chemical, and structural support necessary for culturing a cell.
The term “derived from,” when referring to cells or a biological sample, indicates that the cell or sample was obtained from the stated source at some point in time. For example, a cell derived from an individual can represent a primary cell obtained directly from the individual (i.e., unmodified), or can be modified, e.g., by introduction of a recombinant vector, by culturing under particular conditions, or immortalization. In some cases, a cell derived from a given source will undergo cell division and/or differentiation such that the original cell is no longer exists, but the continuing cells will be understood to derive from the same source.
The term “transfection” or “transfecting” is defined as a process of introducing a nucleic acid molecule to a cell using non-viral or viral-based methods. The nucleic acid molecule can be a sequence encoding complete proteins or functional portions thereof. Typically, a nucleic acid vector, comprising the elements necessary for protein expression (e.g., a promoter, transcription start site, etc.). Non-viral methods of transfection include any appropriate transfection method that does not use viral DNA or viral particles as a delivery system to introduce the nucleic acid molecule into the cell. Exemplary non-viral transfection methods include calcium phosphate transfection, liposomal transfection, nucleofection, sonoporation, transfection through heat shock, magnification and electroporation. For viral-based methods, any useful viral vector can be used in the methods described herein. Examples of viral vectors include, but are not limited to retroviral, adenoviral, lentiviral and adeno-associated viral vectors. In some aspects, the nucleic acid molecules are introduced into a cell using a retroviral vector following standard procedures well known in the art.
Expression of a transfected gene can occur transiently or stably in a host cell. During “transient expression” the transfected nucleic acid is not integrated into the host cell genome, and is not transferred to the daughter cell during cell division. Since its expression is restricted to the transfected cell, expression of the gene is lost over time. In contrast, stable expression of a transfected gene can occur when the gene is co-transfected with another gene that confers a selection advantage to the transfected cell. Such a selection advantage may be a resistance towards a certain toxin that is presented to the cell. Expression of a transfected gene can further be accomplished by transposon-mediated insertion into to the host genome. During transposon-mediated insertion, the gene is positioned in a predictable manner between two transposon linker sequences that allow insertion into the host genome as well as subsequent excision.
The terms “inhibitor,” “repressor” or “antagonist” or “downregulator” interchangeably refer to a substance that results in a detectably lower expression or activity level as compared to a control. The inhibited expression or activity can be 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90% or less than that in a control. In certain instances, the inhibition is 1.5-fold, 2-fold, 3-fold, 4-fold, 5-fold, 10-fold, or more in comparison to a control.
The terms “therapy,” “treatment,” and “amelioration” refer to any reduction in the severity of symptoms, e.g., of a neurodegenerative disorder or neuronal injury. As used herein, the terms “treat” and “prevent” are not intended to be absolute terms. Treatment can refer to any delay in onset, amelioration of symptoms, improvement in patient survival, increase in survival time or rate, etc. The effect of treatment can be compared to an individual or pool of individuals not receiving the treatment, or to the same patient prior to treatment or at a different time during treatment. In some aspects, the severity of disease is reduced by at least 10%, as compared, e.g., to the individual before administration or to a control individual not undergoing treatment. In some aspects the severity of disease is reduced by at least 25%, 50%, 75%, 80%, or 90%, or in some cases, no longer detectable using standard diagnostic techniques.
The term “therapeutically effective amount,” as used herein, refers to that amount of the therapeutic agent sufficient to ameliorate a given disorder or symptoms. For example, for the given parameter, a therapeutically effective amount will show an increase or decrease of at least 5%, 10%, 15%, 20%, 25%, 40%, 50%, 60%, 75%, 80%, 90%, or at least 100%. Therapeutic efficacy can also be expressed as “-fold” increase or decrease. For example, a therapeutically effective amount can have at least a 1.2-fold, 1.5-fold, 2-fold, 5-fold, or more effect over a control.
“Subject,” “patient,” “individual in need of treatment” and like terms are used interchangeably and refer to, except where indicated, mammals such as humans and non-human primates, as well as rabbits, rats, mice, goats, pigs, and other mammalian species. The term does not necessarily indicate that the subject has been diagnosed with a particular disease, but typically refers to an individual under medical supervision.

Methods of Modulating Cardiomyocyte Proliferation

Provided herein are methods of modulating proliferation of a cardiomyocyte using micro RNA modulators. A micro RNA modulator as used herein refers to an agent capable of modulating the level of expression of a micro RNA (e.g. let-7a, mir 100). In some embodiments, the micro RNA modulator is encoded by a nucleic acid. In other embodiments, the micro RNA modulator is a small molecule (e.g. a chemical compound, synthetic micro RNA molecule). In some embodiments, the micro RNA modulator decreases the level of expression of a micro RNA compared to the level of expression in the absence of the micro RNA modulator. Where the micro RNA modulator decreases the level of expression of a micro RNA relative to the absence of the modulator, the micro RNA modulator is an antagonist of said micro RNA. In other embodiments, the micro RNA modulator increases the level expression of a micro RNA compared to the level of expression in the absence of the micro RNA modulator. Where the micro RNA modulator increases the level of expression of a micro RNA relative to the absence of the modulator, the micro RNA modulator is an agonist of the micro RNA.
In one aspect, a method of modulating proliferation of a cardiomyocyte is provided. The method includes (i) transfecting a cardiomyocyte with a nucleic acid encoding a micro RNA modulator, thereby forming a transfected cardiomyocyte; and (ii) allowing the transfected cardiomyocyte to divide, thereby modulating proliferation of the cardiomyocyte. In some embodiments, the nucleic acid is a lentiviral vector. In embodiments, the nucleic acid includes a nucleic acid sequence as set forth in SEQ ID NO:1124 or SEQ ID NO:1125. In some embodiments, the nucleic acid is a lentiviral vector. In embodiments, the nucleic acid includes a nucleic acid sequence as set forth in SEQ ID NO:1124 and SEQ ID NO:1125. In embodiments, the nucleic acid includes a nucleic acid sequence as set forth in SEQ ID NO:1124. In embodiments, the nucleic acid includes a nucleic acid sequence as set forth in SEQ ID NO:1125. In embodiments, the nucleic acid has the sequence as set forth in SEQ ID NO:1124 or SEQ ID NO:1125. In embodiments, the nucleic acid has the sequence as set forth in SEQ ID NO:1124. In embodiments, the nucleic acid has the sequence as set forth in SEQ ID NO:1125.
In other embodiments, the micro RNA modulator is an antagonist of a mir 99 micro RNA, a let-7a micro RNA, a mir 100 micro RNA, a mir 4458 micro RNA, a mir 4500 micro RNA or a mir 89 micro RNA. In some embodiments, the proliferation of the cardiomyocyte is increased compared to a control cardiomyocyte lacking the nucleic acid encoding said RNA modulator. In some embodiments, the micro RNA modulator is an agonist of a mir 99 micro RNA, a let-7a micro RNA, a mir 100 micro RNA, a mir 4458 micro RNA, a mir 4500 micro RNA or a mir 89 micro RNA.
In another aspect, a method of modulating proliferation of a cardiomyocyte is provided. The method includes (i) contacting a cardiomyocyte with a small molecule, thereby forming a treated cardiomyocyte; and (ii) allowing the treated cardiomyocyte to divide, thereby modulating proliferation of the cardiomyocyte. In some embodiments, the proliferation of the cardiomyocyte is increased compared to a control cardiomyocyte lacking the small molecule. In some further embodiment, the small molecule modulates expression of a mir 99 micro RNA-regulated protein, a let-7a micro RNA-regulated protein, a mir 100 micro RNA-regulated protein, a mir 4458 micro RNA-regulated protein, a mir 4500 micro RNA-regulated protein or a mir 89 micro RNA-regulated protein. In other embodiments, the small molecule is a chemical compound. In some embodiments, the small molecule is a synthetic micro RNA molecule. In embodiments, the synthetic micro RNA molecule includes a nucleic acid sequence as set forth in SEQ ID NO:1124 or SEQ ID NO:1125. In embodiments, the synthetic micro RNA molecule includes a nucleic acid sequence as set forth in SEQ ID NO:1124 and SEQ ID NO:1125. In embodiments, the synthetic micro RNA molecule includes a nucleic acid sequence as set forth in SEQ ID NO:1124. In embodiments, the synthetic micro RNA molecule includes a nucleic acid sequence as set forth in SEQ ID NO:1125. In embodiments, the synthetic micro RNA molecule has a nucleic acid sequence as set forth in SEQ ID NO:1124. In embodiments, the synthetic micro RNA molecule has a nucleic acid sequence as set forth in SEQ ID NO:1125.
In other embodiments, the proliferation of the cardiomyocyte is increased compared to a control cardiomyocyte lacking the synthetic micro RNA molecule. In some embodiments, the synthetic micro RNA molecule is an antagonist of a mir 99 micro RNA, a let-7a micro RNA, a mir 100 micro RNA, a mir 4458 micro RNA, a mir 4500 micro RNA or a mir 89 micro RNA. In other embodiments, the synthetic micro RNA molecule is an agonist of a mir 99 micro RNA, a let-7a micro RNA, a mir 100 micro RNA, a mir 4458 micro RNA, a mir 4500 micro RNA or a mir 89 micro RNA.
In another aspect, a method of treating myocardial infarction in a subject in need thereof is provided. The method includes administering to the subject a therapeutically effective amount of a nucleic acid encoding a micro RNA modulator, wherein the RNA modulator increases cardiomyocyte proliferation thereby treating the myocardial infarction. In some embodiments, the micro RNA modulator is an antagonist of a mir 99 micro RNA, a let-7a micro RNA, a mir 100 micro RNA, a mir 4458 micro RNA, a mir 4500 micro RNA or a mir 89 micro RNA. In embodiments, the micro RNA modulator is an antagonist of a mir 99 micro RNA and an antagonist of a let-7a micro RNA. In embodiments, the nucleic acid includes a nucleic acid sequence as set forth in SEQ ID NO:1124 or SEQ ID NO:1125. In embodiments, the nucleic acid includes a nucleic acid sequence as set forth in SEQ ID NO:1124. In embodiments, the nucleic acid includes a nucleic acid sequence as set forth in SEQ ID NO:1125. In embodiments, the nucleic acid includes a nucleic acid sequence as set forth in SEQ ID NO:1124 and SEQ ID NO:1125.
In embodiments, the method includes administering to the subject a therapeutically effective amount of a first nucleic acid and a second nucleic acid, wherein the first nucleic acid encodes an antagonist of a mir 99 micro RNA and the second nucleic acid encodes an antagonist of a let-7a micro RNA. In embodiments, the administering to the subject a therapeutically effective amount of a nucleic acid includes administering a first nucleic acid and a second nucleic acid, wherein the first nucleic acid encodes an antagonist of a mir 99 micro RNA and the second nucleic acid encodes an antagonist of a let-7a micro RNA.
In another aspect, a method of treating myocardial infarction in a subject in need thereof is provided. The method includes administering to the subject a therapeutically effective amount of a nucleic acid encoding an antagonist of a mir 99 micro RNA and a nucleic acid encoding an antagonist of a let-7a micro RNA, thereby treating the myocardial infarction.
In another aspect, a method of treating myocardial infarction in a subject in need thereof is provided. The method includes administering to the subject a therapeutically effective amount of a small molecule, wherein the small molecule increases cardiomyocyte proliferation thereby treating the myocardial infarction. In some embodiments, the small molecule modulates expression of a mir 99 micro RNA-regulated protein, a let-7a micro RNA-regulated protein, a mir 100 micro RNA-regulated protein, a mir 4458 micro RNA-regulated protein, a mir 4500 micro RNA-regulated protein or a mir 89 micro RNA-regulated protein. In embodiments, the small molecule modulates expression of a mir 99 micro RNA-regulated protein and a let-7a micro RNA-regulated protein. In some other embodiments, the small molecule is a chemical compound. In other embodiments, the small molecule is a synthetic micro RNA molecule. In embodiments, the synthetic micro RNA molecule includes a nucleic acid sequence as set forth by SEQ ID NO:1124 or SEQ ID NO:1125. In embodiments, the synthetic micro RNA molecule includes a nucleic acid sequence as set forth by SEQ ID NO:1124 and SEQ ID NO:1125. In embodiments, the synthetic micro RNA molecule includes a nucleic acid sequence as set forth by SEQ ID NO:1124. In embodiments, the synthetic micro RNA molecule includes a nucleic acid sequence as set forth by SEQ ID NO:1125. In some embodiments, the synthetic micro RNA molecule is an antagonist of a mir 99 micro RNA, a let-7a micro RNA, a mir 100 micro RNA, a mir 4458 micro RNA, a mir 4500 micro RNA or a mir 89 micro RNA.

EXAMPLES

Heart failure remains one of the leading causes of mortality in the developed world. Whereas the mammalian heart is endowed with certain regenerative potential, endogenous cardiomyocyte proliferation is insufficient for functional heart repair upon injury. Interestingly, non-mammalian vertebrates, such as the zebrafish, can regenerate the damaged heart by inducing cardiomyocyte dedifferentiation and proliferation. By screening regenerating zebrafish hearts Applicants identified miR-99/100 down-regulation as a key process driving cardiomyocyte dedifferentiation. Experimental down-regulation of miR-99/100 in primary adult murine and human cardiomyocytes led to an increase in the number of proliferating cardiomyocytes. AAV-mediated in vivo down-regulation of miR-99/100 after acute myocardial injury in mice induced mature cardiomyocyte proliferation, diminished infarct size and improved heart function. Applicants' study unveils conserved regenerative mechanisms between zebrafish and mammalian cardiomyocytes and represents a proof-of-concept on the suitability of activating pro-regenerative responses for healing the diseased mammalian heart.
Cardiovascular disease remains the leading cause of mortality in the developed world. Attempts at developing curative strategies have mainly focused on the activation of endogenous cardiac progenitor cells and the transplantation of in vitro-derived cardiomyocytes (A. Aguirre et al., Cell Stem Cell 12, 275-284 (2013); S. R. Braam et al., Trends in pharmacological sciences 30, 536-45 (2009)). More recently, in vivo reprogramming strategies have emerged as potential treatments for heart failure (L. Qian et al., Nature (2012), doi:10.1038/nature11044; K. Song et al., Nature 485, 599-604 (2012) A. Eulalio et al., Nature (2012), doi:10.1038/nature11739). Along this line, a recent report by Porrello et al. has highlighted a remarkable regenerative capacity in neonatal murine hearts upon injury (E. R. Porrello et al., Science (New York, N.Y.) 331, 1078-80 (2011)). Although adult mammalian cardiomyocytes retain a certain ability to proliferate (S. E. Senyo et al., Nature, 2-6 (2012)), endogenous regenerative responses during adulthood are largely insufficient for replenishing the lost cardiac tissue. Noticeably, heart repair can be induced upon manipulation of miRNA pathways identified as drivers of cardiomyocyte proliferation in neonatal murine models (A. Eulalio et al., Nature (2012), doi:10.1038/nature11739L; E. R. Porrello et al., Circulation research 109, 670-9 (2011)). This may suggest that the mechanisms underlying heart regeneration at birth are still present, yet dormant and/or repressed, in adult murine hearts. Other vertebrates, such as the zebrafish, are able, throughout their entire lifetime, to activate endogenous regenerative responses that lead to complete cardiomyocyte-mediated heart regeneration similar to that observed in neonatal mice (R. Zhang et al., Nature (2013), doi:10.1038/nature12322; K. D. Poss et al., Science (New York, N.Y.) 298, 2188-90 (2002); A. Raya et al., Proceedings of the National Academy of Sciences of the United States of America 100 Suppl, 11889-95 (2003); C. Jopling et al., Nature 464, 606-9 (2010); K. Kikuchi et al., Nature 464, 601-5 (2010)). Together, these observations led us to hypothesize on the existence of conserved pro-regenerative pathways between zebrafish and mammals (A. W. Seifert et al., Nature 489, 561-5 (2012)), and if present, whether they could be altered to drive terminally differentiated mammalian cardiomyocytes to a regeneration-competent state.
To first elucidate the presence of conserved regulatory pathways underlying regeneration Applicants decided to focus on microRNAs promoting cardiomyocyte dedifferentiation in the zebrafish. Expression of 90 microRNAs was significantly changed 3 days post amputation (dpa) of the ventricular apex (FIG. 5A). Bioinformatic analysis of signaling pathways and GO processes indicated significant enrichment in proliferation pathways, as well as processes related to chromatin remodeling (S. L. Paige et al., Cell 151, 221-32 (2012)), morphogenesis and kinase activity (FIG. 5B). Interestingly, two well-defined down-regulated microRNA clusters (miR-99/Let-7a and miR-100/Let-7c) were highly conserved in sequence and genomic organization across different vertebrates (FIG. 5C). Putative protein targets were also shared between zebrafish and mammals (Table S1 and Table S2). Further expression analysis demonstrated a significant down-regulation of miR-99/100 and Let-7a/c during the early regenerative stages (3-7 dpa) (FIG. 1A) in agreement with previous reports (C. Jopling et al., Nature 464, 606-9 (2010)). Noticeably, miR-99/100 and Let-7a/c expression (data not shown) was high and confined to cardiomyocytes in uninjured hearts, and almost undetectable upon injury (FIGS. 1, B and C and FIG. 6). microRNA target prediction highlighted two proteins specifically expressed in the regenerating zebrafish heart, Fntβ (beta subunit of farnesyl-transferase) and Smarca5 (SWI/SNF-related matrix associated actin-dependent regulator of chromatin subfamily a, member 5) (FIGS. 1, B to D and FIG. 6). Binding experiments confirmed miRNA targeting of the 3′UTRs in both human and zebrafish Fntβ and Smarca5 (a C. Mueller et al., 2H., Oncogene, 1-9 (2012)) (FIG. 7). Accompanying Fntβ up-regulation, the structural subunit of fnt (Fntα) was also expressed during heart regeneration (FIG. 8). Chemical inhibition of fnt with the specific antagonist tipifarnib significantly impaired heart regeneration by decreasing the number of proliferating cardiomyocytes (FIG. 1, E to G). Taken together, these data suggest that targets downstream of miR-99/100 play a functional role during heart regeneration.
Since regeneration might be considered to a large extent redolent of development (J. P. Brockes, A. Kumar, Annual review of cell and developmental biology 24, 525-49 (2008); J. L. Whited, C. J. Tabin, 2-4 (2010); D. Knapp, E. M. Tanaka, Current Opinion in Genetics & Development (2012), doi:10.1016/j.gde.2012.09.006) Applicants next decided to investigate the role of miR-99/100 and their target proteins Fntβ and Smarca5 during zebrafish heart development and maturation. qRT-PCR and immunofluorescence analyses demonstrated low levels of miR-99/100 expression during early heart development concomitantly with high levels of Smarca5 and Fntβ (FIG. 9). Functional analyses by injection of miR-99/100 mimics, and/or fntβ/smarca5 translation-blocking morpholinos in one-cell stage cm1c2:GFP transgenic fish embryos resulted in a significantly reduced ventricle size in spite of the presence of apparently normal heart anatomical structures (ventricle, atrium, valve) (FIG. 1, H to J). To determine if cardiomyocytes showing up-regulation of Fntβ and Smarca5 progress into a proliferative state, Applicants analyzed the expression of nuclear proliferative markers at different time points post-amputation (FIG. 2, A to D). In all cases high levels of Fntβ and Smarca5 correlated with PCNA and/or H3P expression in the nucleus. Concomitantly, disorganized sarcomeric structures were particularly evident at 7 dpa (FIGS. 2, A and B). miR-99/100 expression levels, and their respective protein targets, returned to basal levels when regeneration of the ventricle was mostly complete at 30 dpa (FIGS. 10, A and B). Next, Applicants designed a series of in vivo experiments to exogenously manipulate miR-99/100 levels with mimics and antagomiRs in adult regenerating animals (FIGS. 10, C and D). Intra-cardiac injection of miR-99/100 mimics efficiently blocked the regenerative response in all animals tested (FIGS. 2, E and F) and BrdU incorporation confirmed that cardiomyocyte proliferation was significantly disrupted in mimic-treated animals (FIG. 2G). Conversely, microRNA inhibition of size-matched sibling fish led to significantly enlarged hearts (FIGS. 2, H and I). Histological analysis indicated cardiomyocyte proliferation in the absence of cardiac hypertrophy (FIG. 2J), suggesting hyperplasia as the underlying mechanism of action of miRNA-99/100. Applicants next decided to study the downstream signaling mechanisms of miR-99/100 and Let-7a/c. Applicants detected increased farnesylation as a consequence of fnt activity in regenerating hearts at 3 and 7 dpa (FIG. 11). Ras family proteins, targets of fnt, appeared up-regulated and preferentially located to the cell membrane (FIG. 12, A to C). Similarly, c-Myc, a transcription factor essential for cellular proliferation downstream of this pathway, was significantly up-regulated and localized to the cell nucleus in dedifferentiating cardiomyocytes (FIG. 12, D to F). Interestingly, Cbx5 and Cbx3a, chromatin-remodeling proteins critical in proliferating cardiomyocytes (J. K. Takeuchi et al., Nature communications 2, 187 (2011); N. Collins et al., Nature genetics 32, 627-32 (2002); S. H. Kwon, J. L. Workman, BioEssays: news and reviews in molecular, cellular and developmental biology 33, 280-9 (2011)), demonstrated enhanced expression at 7 dpa, when Smarca5 levels in the nucleus were highest (FIG. 13). Taken together, Applicants' results indicate that miR-99/100 down-regulation plays a role, possibly by chromatin remodeling, in the dedifferentiation process that leads to zebrafish heart regeneration (FIG. 14).
In light of the evolutionary conservation of their structures and downstream signaling pathways, Applicants wondered whether microRNA-99/100 and Let-7a/c functions would be similar between mammals and zebrafish. To this end, Applicants first analyzed microRNA-99/100 and FNTβ/SMARCA5 expression in developing and adult murine hearts. qRT-PCR and immunofluorescence analyses highlighted a progressive up-regulation of microRNA-99/100 paralleling cardiac maturation and FNTβ/SMARCA5 down-regulation (FIGS. 3, A and B and FIG. 15). Analyses of human cardiomyocytes at progressive differentiation stages, including adult heart samples, demonstrated a peak in miR-99/100 expression in adult mature human cardiomyocytes, a point at which FNTβ/SMARCA5 expression was undetectable (FIGS. 3, C and D). hESC-derived immature proliferative cardiomyocytes (hiCM) expressed GATA4 (a cardiac progenitor marker), FNTβ, SMARCA5 and intermediate-low levels of the identified miRNAs (FIG. 16) resembling a regenerative, proliferative state as described before (K. Kikuchi et al., Nature 464, 601-5 (2010), C. Jopling et al., Nature reviews. Molecular cell biology 12, 79-89 (2011)).
Applicants next sought to evaluate the effects of microRNA down-regulation in adult murine cardiomyocytes. Seven days after shRNA-mediated microRNA silencing significant up-regulation of SMARCA5 and FNTβ was observed accompanied by an increased amount of cardiomyocytes with disorganized sarcomeric structures and immature morphology (FIGS. 3, E and F and FIG. 17, A to C). Further analysis demonstrated enhanced proliferation paralleling GATA4 and PCNA re-expression (FIG. 3, E to G and FIG. 17). These effects were more pronounced when miR-99/100 and Let-7a/c were simultaneously blocked (FIG. 3G and FIG. 17). Similarly, microRNA silencing in human cardiomyocytes resulted in increased proliferation and higher numbers of beating colonies (FIGS. 3, G and H and Videos 1 to 5). Functional analyses demonstrated minimal changes in the beating rate before and after anti-miR delivery (FIG. 17D). Indicative of cardiomyocyte specificity, microRNA down-regulation did not affect proliferation or FNTβ/SMARCA5 expression in human fibroblasts or vascular cells (FIG. 18). Organotypic cultures of adult murine hearts, a setting more closely resembling physiological conditions (M. Brandenburger et al., Cardiovascular research 93, 50-9 (2012)), demonstrated low to undetectable levels of FNTβ/SMARCA5 (FIG. 19). Delivery of anti-miR-99/100 and/or anti-miR-Let-7 to murine heart organotypic slices resulted in cardiomyocyte proliferation as demonstrated by down-regulation of MyHC as well as re-expression of GATA4 and H3 phosporylation (FIG. 19). Ultrastructural electron microscopy analysis confirmed cytoskeletal disassembly (FIG. 4A). Further supporting these observations, Connexin 43 (Cx43), an essential component of coupling GAP junctions in cardiomyocytes, was profoundly down-regulated (FIG. 19). Next, Applicants mimicked tissue ischemia and heart damage in organotypic slices to determine the effects of the treatment upon injury (FIG. 20A). Control organotypic slices developed necrotic areas accompanied by marginal proliferation under hypoxic conditions whereas anti-microRNA delivery resulted in reduced necrosis (FIG. 20B to D) and the appearance of proliferative cardiomyocyte populations (FIG. 4B and FIG. 20D).
Lastly, Applicants decided to test the efficacy of anti-microRNA delivery for the induction of regeneration in a murine model of myocardial infarction. Following LAD artery ligation, anti-miR-99/100 and anti-Let-7 were administered by injection of serotype 9 adeno-associated viral (AAV) particles, specifically targeting the cardiomyocyte population, in the periphery of the infarcted area. 18 days after treatment, both ejection fraction and fractional shortening significantly improved in the treated group (FIG. 4, C to F). Reduced fibrotic scarring and infarct sizes were readily observed three weeks after LAD artery ligation, indicative of an underlying regenerative response (FIG. 4G). Treated animals exhibited increased numbers of FNTβ/SMARCA5 positive cardiomyocytes, as well as a marked increase of cardiomyoctes re-expressing GATA4. PCNA and H3P staining demonstrated increased DNA synthesis and cardiomyocyte mitosis (FIG. 4, H to L). Of note, the number of mitotic cardiomyocytes was higher in areas of trabeculated muscle as opposed to the myocardium proper. Taken together, all these observations indicate that anti-miR delivery in adult murine cardiomyocytes suffices for the induction of a pro-regenerative proliferative response towards repairing a damaged heart.
These observations constitute a proof-of-concept on how animal models naturally capable of regeneration can be used for the identification of regenerative factors that may, subsequently, be applied to mammals. Experimental manipulation of conserved microRNAs unveiled during adult zebrafish heart regeneration led to similar responses in mice after heart infarction (replenishment of the lost cardiac tissue and inhibition of scar formation). In vivo activation of conserved cardiac regenerative responses may help to circumvent many of the problems associated with heart cell transplantation as well as those associated with reprogramming technologies (A. Aguirre et al., Cell Stem Cell 12, 275-284 (2013), M. a. Laflamme, C. E. Murry, Nature 473, 326-335 (2011)), serving as an additional tool to the clinical armamentarium of regenerative medicine towards the treatment of human heart disease (K. R. Chien et al., Journal of molecular and cellular cardiology 53, 311-3 (2012)).

Experimental Procedures

Detailed experimental procedures can be found in Supplementary information.
Animals. Wild-type zebrafish (AB) and cm1c2:GFP were maintained at 28.5° C. by standard methods, unless otherwise indicated. All protocols were previously approved and performed under institutional guidelines.
Culture and isolation of adult mouse ventricular myocytes. Wild-type mice (C57B6/J) were sacrificed and hearts were quickly recovered and washed with ice-cold Ca²⁺-free ModifedTyrode's Solution (MTS). Ventricles were dissected from the rest of the heart and subjected to enzymatic digestion (Liberase DH, Roche) for 10-15 min in a spinner flask at 37 C under continuous agitation. Afterwards cells were pelleted by short centrifugation, resuspended in KB solution and cardiomyocytes were left to sediment by gravity, thus greatly reducing the presence of other contaminating cell types. Calcium was restore to 1 mM in a step-wise fashion in three gradual steps and subsequently cardiomyocytes were centrifuged, resuspended in culture medium (IMDM 5%, 1% Pen/Strep, 0.1 ng/ml FGFb, 1 ng/ml TGF-β3) and seeded in laminin-coated tissue-culture plates. Cells were kept in culture for 1 week.
Lentiviral and AAV constructs. Anti-miR constructs, miRZip-99/100 and miRZip-let7 (SBI), were used according to the manufacturer instructions. As respective controls, the anti-miRs were removed from the parent vector by digesting with BamH1 and EcoR1, end filled and re-ligated. Lentiviruses were packaged by transfecting in 293T cells followed by spinfection in the respective mouse or human ES derived cardiomyocytes. AAVs were generated as described before (Eulalio et al, 2012). Briefly, the antimiR constructs contained in the miRZip vectors were excised and ligated into pZacf-U6-luc-ZsGreen. Serotype 9 AAVs were packaged by transfection of 293T cells with the appropriate plasmids.
Organotypic heart slice culture. Mice ventricles were washed in cold Modified Tyrode's Solution, embedded in 4% low melting point agarose and immediately cut into 300 μm slices using a vibratome (Leica). Heart slices were then maintained in complete IMDM 5%, 1% Pent/Strep in 12-well plates at the medium-air interface using 0.4 μm membrane transwells (Corning) at 37 C in a 5% CO₂incubator. For experimental hypoxia-like conditions, slices were kept in a hypoxia chamber incubator for 4 hours at 37 C, 5% O₂. Lentiviral transduction was performed by immersion of the slices in virus-containing medium for 24 h.
Myocardial infarction. Myocardial infarction was induced CD1 mice (8-12 weeks old) by permanent left anterior descending (LAD) coronary artery ligation. Briefly, mice were anesthetized with an injection of ketamine and xylazine, intubated and placed on a rodent ventilator. Body temperature was maintained at 37° C. on a heating pad. After removing the pericardium, a descending branch of the LAD coronary artery was visualized with a stereomicroscope and occluded with a nylon suture. Ligation was confirmed by the whitening of a region of the left ventricle. Recombinant AAV vectors, at a dose of 10¹¹viral genome particles per animal, were injected immediately after LAD ligation into the myocardium bordering the infarct zone (single injection), using an insulin syringe with incorporated 30-gauge needle. Three groups of animals were studied, receiving AAV9-control (shRNA-Luc), AAV9-antimiR-99/100 or AAV9-anti-Let-7a/c. The chest was closed, and the animals moved to a prone position until the occurrence of spontaneous breathing. BrdU was administered intraperitoneally (500 μg per animal) every 2 days, for a period of ten days. Echocardiography analysis was performed at days 12, 30 and 60 after infarction, as described below, and hearts were collected at 12 (n=6 animals per group) and 60 (n=10 animals per group) days after infarction.
Echocardiography analysis. To evaluate left ventricular function and dimensions, transthoracic two-dimensional echocardiography was performed on mice sedated with 5% isoflurane at 12, 30 and 60 days after myocardial infarction, using a Visual Sonics Vevo 770 Ultrasound (Visual Sonics) equipped with a 30-MHz linear array transducer. M-mode tracings in parasternal short axis view were used to measure left ventricular anterior and posterior wall thickness and left ventricular internal diameter at end-systole and end-diastole, which were used to calculate left ventricular fractional shortening and ejection fraction.
Heart collection and histological analysis. At the end of the studies, animals were anaesthetized with 5% isoflurane and then killed by injection of 10% KCl, to stop the heart at diastole. The heart was excised, briefly washed in PBS, weighted, fixed in 10% formalin at room temperature, embedded in paraffin and further processed for histology or immunofluorescence. Haematoxylin—eosin and Masson's trichrome staining were performed according to standard procedures, and analysed for regular morphology and extent of fibrosis. Infarct size was measured as the percentage of the total left ventricular area showing fibrosis.
Zebrafish heart amputation. Adult fish were anaesthetized in 0.4% Tricaine and secured, ventral side uppermost, in a slotted sponge. Watchmaker forceps were used to remove the surface scales and penetrate the skin, muscle and pericardial sac. Once exposed, the ventricle was gently pulled at the apex and cut with iridectomy scissors. After surgery, fish were immediately returned to system water.
Cryosectioning. At the specified time points, hearts were removed, washed in PBS-EDTA 0.4% and fixed for 20 min in 4% paraformaldehyde at 4° C. Afterwards, they were washed several times in PBS, equilibrated in 30% sucrose, and then frozen for cryosectioning. 10 tm slices were obtained with a cryostat (Leica).
Real time RT-PCR. For RNA, tissue was obtained from adult heart ventricles from different time points and conditions, extensively washed in PBS-EDTA 0.4% to remove blood, and then mechanically homogenized and processed using RNeasy kit (Qiagen) as per manufacturer's instructions. RT and PCR were performed using Quantitect Reverse Transcription Kit (Qiagen) and Quantitect Primers for the following genes: Fntb, Fnta, Smarca5, myc-a, myc-b, H-rasa, H-rasb, N-ras, K-ras, tnnt2. For miRNAs, small RNA (<200 pb) was obtained employing the miRNeasy mini kit (Qiagen) using the same procedure as before. RT and PCR reactions were carried out employing miRCURY LNA RT and PCR kits (Exiqon) and stem-loop LNA primers (Exiqon).
MicroRNA microarrays. RNA was obtained as for PCR applications. GenechipmiRNA 2.0 microarrays were purchased from Affymetrix and small RNA labeling was performed using FlashTag HSR labeling kit (Genesphere). 200 ng of small RNA was polyA-tailed and biotin conjugated. After labelling, RNA was hybridized using GeneChip reagents (Affymetrix) and protocols as indicated by the manufacturer. The chip contains hybridization probes for the miRbase v15 annotations, including 248 zebrafish miRNAs. MicroRNA data was analyzed by using the R package.
Bioinformatic analysis of miRNA targets. Signaling pathways and downstream target prediction related to the identified miRNAs were determined by using DIANA, Miranda and TargetScan. Gene ontology analysis was performed with DAVID software.
Fluorescence in situ hybridization. 10 μm heart slices were further fixed in 4% PFA for 10 min at room temperature, washed in PBS and acetylated for 10 min in acetylation solution. After washing in PBS, samples were treated with proteinase K, prehybridized for 4 h and hybrydized overnight at the appropriate temperature with LNA DIG-labeled probes for the corresponding miRNAs (Exiqon). The next day slides were washed and immunolabeled with anti-DIG-alkaline phosphatase antibodies (1:2,000) and antibodies against cardiomyocytic proteins of interest (1:100) overnight at 4° C. Secondary antibody incubation was performed as for immunofluorescence experiments. Alkaline phosphatase activity was detected by incubating samples in a Fast Red solution (Dako) for 2 hours. Samples were then washed, mounted in Vecta-shield and imaged in a confocal microscope. Fast Red fluorescence was detected with Cy3 settings.
Immunofluorescence. Tissue slices were fixed for 15 min in 4% paraformaldehyde, washed in PBS-gly 0.3 M, and blocked in PBS-10% donkey serum, 0.5% TX-100, 0.5% BSA for 1 hour. Primary antibodies were diluted at the appropriate concentrations in PBS-1% donkey serum, 0.5% TX-100, 0.5% BSA and incubated overnight. After washing, slices were incubated overnight with secondary antibodies, washed and mounted in Vecta-shield. Antibodies employed are listed in table S3.
Cell culture. COS7 cells were maintained in DMEM (high glucose) supplemented with 10% FBS, L-Glutamine and non-essential amino acids (Invitrogen). Human ES cells, H1 and H9 (WA1 and WA9, WiCell), were cultured in chemically defined hES/hiPS growth media, mTeSR1 on growth factor reduced matrigel (BD biosciences) coated plates. Briefly, 70-80% confluent hES/iPS cells were treated with dispase (Invitrogen) for 7 minutes at 37° C. and the colonies were dispersed to small clusters and lifted carefully using a 5 ml glass pipette, at a ratio of ˜1:4.
Culture and isolation of adult mouse ventricular myocytes. Wild-type mice (C57B6/J) were sacrificed and hearts were quickly recovered and washed with ice-cold Ca²⁺-free ModifedTyrode's Solution (MTS). Ventricles were dissected from the rest of the heart and subjected to enzymatic digestion (Liberase DH, Roche) for 10-15 min in a spinner flask at 37 C under continuous agitation. Afterwards cells were pelleted by short centrifugation, resuspended in KB solution and cardiomyocytes were left to sediment by gravity, thus greatly reducing the presence of other contaminating cell types. Calcium was restore to 1 mM in a step-wise fashion in three gradual steps and subsequently cardiomyocytes were centrifuged, resuspended in culture medium (IMDM 5%, 1% Pen/Strep, 0.1 ng/ml FGFb, 1 ng/ml TGF-β3) and seeded in laminin-coated tissue-culture plates. Cells were kept in culture for 1 week.
Differentiation of Human ES cells to immature cardiomyocytes. Human ES cells grown on matrigel dots (BD biosciences) were carefully dissociated using dispase and were plated on low attachment plates in EB media (IMDM, 20% FBS, 2.25 nM L-Glutamine and non-essential aminoacids). After 6 days of suspension in culture, the EBs were seeded on gelatin-coated plates in EB media. Spontaneously beating EBs were manually picked and used for further analysis. For directed differentiation, human ES cells grown in mTeSR on matrigel coated plates were treated with 12 μM GSK3β inhibitor CHIR 99021 (Stemgent) in cardiomyocyte differentiation base media (RPMI 1640 supplemented with 125 μg/ml human holo-transferrin (Sigma-Aldrich)) for 24 hours, followed by 24 hour of rest in the base media. On day 3, the cells were treated with 5 μM WNT inhibitor, IWP4 (Stemgent) for 48 hours, followed by treatment with Cardiac differentiation base media supplemented with 20 μg/ml human Insulin (SAFC) until colonies started beating.
Lentiviral constructs. Anti-miR constructs, miRZip-99/100 and miRZip-let7 (SBI), were used according to the manufacturer instructions. As respective controls, the anti-miRs were removed from the parent vector by digesting with BamH1 and EcoR1, end filled and re-ligated. Lentiviruses were packaged by transfecting in 293T cells followed by spinfection in the respective mouse or human ES derived cardiomyocytes.
Luciferase constructs and microRNA binding validation. 3′ UTR of human and zebrafish FNTB and SMARCA5 were amplified with the indicated primers using genomic DNA as a template and were cloned into PGL3 vector (Promega) at the Xho1 site downstream of luciferase gene. COS7 cells (seeded at 3×10⁴cells per well of a 12 well plate and grown for 24 hours) were transfected with 50 ng each of indicated luciferase reporter vectors, pRL TK (Renilla luciferase control vector, Promega) either in the presence or absence of 20 nM or 40 nM of double stranded DNA oligonucleotide mimics of miR-99 or miR-100 (Dharmacon) using Lipofectamine (Invitrogen) following manufacturer's protocol. 12-16 hours post-transfection, cells were lysed using passive lysis buffer (Promega). Luminescent signals arising from the cell lysates obtained 12 hours post transfection of COST cells with appropriate luciferase constructs were measured using the Dual Luciferase assay system (Promega) in a Synergy H1 hybrid reader (BioTek). The relative luminescence intensity of each sample was calculated after normalization with corresponding Renilla luciferase activity, and were represented as % values compared to the corresponding sample without the miR mimic.
Confocal microscopy. Samples were imaged using a Zeiss L710 confocal microscope. For every sample, at least two different fields were examined at two different magnifications (using a 20× objective and a 63× oil-immersion objective). Z-stacks were obtained for further analysis and 3D reconstruction. For intensity comparison purposes, images were taken with the same settings (pinhole size, laser intensity, etc).
Organotypic heart slice culture. Mice ventricles were washed in cold Modified Tyrode's Solution, embedded in 4% low melting point agarose and immediately cut into 300 μm slices using a vibratome (Leica). Heart slices were then maintained in complete IMDM 5%, 1% Pent/Strep in 12-well plates at the medium-air interface using 0.4 μm membrane transwells (Corning) at 37 C in a 5% CO₂incubator. For experimental hypoxia-like conditions, slices were kept in a hypoxia chamber incubator for 4 hours at 37 C, 5% O₂. Lentiviral transduction was performed by immersion of the slices in virus-containing medium for 24 h.
Morpholino and microRNA injections in zebrafish embryos. Morpholinos (Gene Tools) were dissolved in water at a 2 mM stock concentration and diluted to a 2 ng/nl working concentration in PBS/phenol red solution. Embryo injections were performed by injecting ˜1 nl morpholino solution at the 1-cell stage using a FemtoJet (Eppendorf). For microRNA mimic injection, a miR-99/100 equimolar mixture at 2 ng/nl in PBS was employed. Morphants were evaluated at 24, 48 and 72 h in a StereoLumar stereoscope (Zeiss).
In vivo microRNA delivery. MicroRNA siRNA mimics without chemical modifications were purchased from Life Technologies, dissolved in nuclease-free water and complexed to jetPEI (10 N/P ratio) for in vivo, intra-cardiac administration. 0.2 μg siRNA was injected per animal every 2 days. To determine the efficiency of the delivery, a control Cy5-labeled siRNA directed against GFP was used in cm1c2:GFP animals. MicroRNA inhibitors against the miR-99/100 family were purchased from Exiqon and used at 0.2 μgsiRNA per animal every 2 days.
Tipifarnib injections. Tipifarnib was dissolved in DMSO at 10 mg/ml and 2 μl were administered by intraperitoneal injection (final concentration 0.02 mg/animal) every 2 days for 14 days. Control animals were administered DMSO.
BrdU labeling. Fish were anaesthetized in 0.4% Tricaine, and 10 μl of a 10 mg/ml solution of BrdU (in PBS) was injected into the abdominal cavity once every 2 days for 14 days. At that point, hearts were removed and fixed overnight in 4% paraformaldehyde at 4° C., washed in PBS, equilibrated in 30% sucrose in PBS and frozen for cryosectioning.
Histology and histomorphometry. Masson's trichrome staining was performed in 10 μm tissue slices by immersion in Bouin's fixative followed by sequential incubation in Weigert'shematoxylin, Acid Fuchsin, phosphotungstic/phosphomolybdic acid, Aniline Blue and acetic acid. After washes, slices were mounted for bright field observation. Histomorphometric measurements were performed with Fiji. Injured areas were quantified in four independent different slices per animal (four animals were used per condition) and normalized to whole tissue area.
Statistical analysis. Results are expressed as mean±SEM. Statistical significance was determined by Student's t-test. Results are representative of at least 3 independent experiments except when otherwise indicated.

TABLE S1

miR-99/100 predicted targets

		SEQ			Total
Target	Representative	ID		Representative	context +	Aggregate	Publication
gene	transcript	NO:	Gene name	miRNA	score	PCT	(s)

FGFR3	NM_000142	1	fibroblast growth	hsa-miR-99a	−0.47	<0.1	2005,
			factor receptor 3				2007,
							2009
IGF1R	NM_000875	2	insulin-like growth	hsa-miR-100	−0.26	<0.1	2007,
			factor 1 receptor				2009
PPP3CA	NM_000944	3	protein phosphatase	hsa-miR-99a	−0.26	<0.1	2009
			3, catalytic subunit,
			alpha isozyme
ZNF197	NM_001024855	4	zinc finger protein	hsa-miR-100	−0.65	<0.1
			197
TTC39A	NM_001080494	5	tetratricopeptide	hsa-miR-99a	−0.55	<0.1	2007,
			repeat domain 39A				2009
NXF1	NM_001081491	6	nuclear RNA export	hsa-miR-100	−0.2	0.11	2009
			factor 1
SMARCA4	NM_001128844	7	SWI/SNF related,	hsa-miR-100	−0.26	0.11
			matrix associated,
			actin dependent
			regulator of
			chromatin, subfamily
			a, member 4
LRRC8B	NM_001134476	8	leucine rich repeat	hsa-miR-99a	−0.29	<0.1
			containing 8 family,
			member B
EIF2C2	NM_001164623	9	eukaryotic translation	hsa-miR-100	−0.4	<0.1	2005,
			initiation factor 2C, 2				2007,
							2009
TMEM135	NM_001168724	10	transmembrane	hsa-miR-100	−0.28	0.11
			protein 135
CLDN11	NM_001185056	11	claudin 11	hsa-miR-100	−0.29	<0.1	2009
BMPR2	NM_001204	12	bone morphogenetic	hsa-miR-100	−0.28	0.11	2005,
			protein receptor, type				2007,
			II (serine/threonine				2009
			kinase)
INSM1	NM_002196	13	insulinoma-	hsa-miR-99a	−0.25	<0.1	2005,
			associated 1				2007
PPP1CB	NM_002709	14	protein phosphatase	hsa-miR-100	−0.31	0.11	2009
			1, catalytic subunit,
			beta isozyme
FZD5	NM_003468	15	frizzled family	hsa-miR-100	−0.3	<0.1	2007,
			receptor 5				2009
SMARCA5	NM_003601	16	SWI/SNF related,	hsa-miR-100	−0.45	<0.1	2005,
			matrix associated,				2007,
			actin dependent				2009
			regulator of
			chromatin, subfamily
			a, member 5
PPFIA3	NM_003660	17	protein tyrosine	hsa-miR-100	−0.3	0.11	2005,
			phosphatase, receptor				2007,
			type, f polypeptide				2009
			(PTPRF), interacting
			protein (liprin), alpha 3
MTOR	NM_004958	18	mechanistic target of	hsa-miR-100	−0.38	<0.1	2005,
			rapamycin				2007,
			(serine/threonine				2009
			kinase)
ST5	NM_005418	19	suppression of	hsa-miR-100	−0.33	<0.1
			tumorigenicity 5
HOXA1	NM_005522	20	homeobox A1	hsa-miR-99a	−0.33	<0.1	2005,
							2007,
							2009
HS3ST3B1	NM_006041	21	heparan sulfate	hsa-miR-99a	−0.59	<0.1	2005,
			(glucosamine) 3-O-				2007,
			sulfotransferase 3B1				2009
HS3ST2	NM_006043	22	heparan sulfate	hsa-miR-100	−0.6	<0.1	2005,
			(glucosamine) 3-O-				2007,
			sulfotransferase 2				2009
ICMT	NM_012405	23	isoprenylcysteine	hsa-miR-99a	−0.15	<0.1	2005,
			carboxyl				2007,
			methyltransferase				2009
BAZ2A	NM_013449	24	bromodomain	hsa-miR-100	−0.46	<0.1	2005,
			adjacent to zinc				2007,
			finger domain, 2A				2009
ZZEF1	NM_015113	25	zinc finger, ZZ-type	hsa-miR-100	−0.42	<0.1	2005,
			with EF-hand				2007,
			domain 1				2009
NIPBL	NM_015384	26	Nipped-B homolog	hsa-miR-99a	−0.3	0.11
			(Drosophila)
PI15	NM_015886	27	peptidase inhibitor	hsa-miR-99a	−0.19	0.11	2009
			15
ZBTB7A	NM_015898	28	zinc finger and BTB	hsa-miR-100	−0.16	0.11	2007,
			domain containing				2009
			7A
PPPDE1	NM_016076	29	PPPDE peptidase	hsa-miR-99a	−0.26	0.11	2009
			domain containing 1
EPDR1	NM_017549	30	ependymin related	hsa-miR-100	−0.69	<0.1	2009
			protein 1 (zebrafish)
RAVER2	NM_018211	31	ribonucleoprotein,	hsa-miR-99a	−0.44	<0.1	2007,
			PTB-binding 2				2009
CYP26B1	NM_019885	32	cytochrome P450,	hsa-miR-100	−0.14	<0.1	2005,
			family 26, subfamily				2007,
			B, polypeptide 1				2009
TAOK1	NM_020791	33	TAO kinase 1	hsa-miR-100	−0.21	<0.1
MBNL1	NM_021038	34	muscleblind-like	hsa-miR-99a	−0.2	<0.1	2005,
			(Drosophila)				2007,
							2009
MTMR3	NM_021090	35	myotubularin related	hsa-miR-99a	−0.18	<0.1	2005,
			protein 3				2007,
							2009
ADCY1	NM_021116	36	adenylate cyclase 1	hsa-miR-99a	−0.4	<0.1	2005,
			(brain)				2007,
							2009
RRAGD	NM_021244	37	Ras-related GTP	hsa-miR-100	−0.35	<0.1
			binding D
TRIB2	NM_021643	38	tribbles homolog 2	hsa-miR-100	−0.31	<0.1	2005,
			(Drosophila)				2007,
							2009
CEP85	NM_022778	39	centrosomal protein	hsa-miR-99a	−0.29	0.11	2007,
			85 kDa				2009
RMND5A	NM_022780	40	required for meiotic	hsa-miR-99a	−0.15	0.11
			nuclear division 5
			homolog A (S. cerevisiae)
THAP2	NM_031435	41	THAP domain	hsa-miR-100	−0.64	0.11	2007,
			containing, apoptosis				2009
			associated protein 2
FZD8	NM_031866	42	frizzled family	hsa-miR-100	−0.34	<0.1	2005,
			receptor 8				2007,
							2009
KBTBD8	NM_032505	43	kelch repeat and	hsa-miR-100	−0.6	<0.1	2007,
			BTB (POZ) domain				2009
			containing 8
SLC44A1	NM_080546	44	solute carrier family	hsa-miR-100	−0.31	<0.1	2007,
			44, member 1				2009
ZNRF2	NM_147128	45	zinc and ring finger 2	hsa-miR-100	−0.24	0.11
ST6GALNAC4	NM_175039	46	ST6 (alpha-N-acetyl-	hsa-miR-99a	−0.56	<0.1
			neuraminyl-2,3-beta-
			galactosyl-1,3)-N-
			acetylgalactosaminide
			alpha-2,6-
			sialyltransferase 4
GRHL1	NM_198182	47	grainyhead-like 1	hsa-miR-99a	−0.3	0.11	2009
			(Drosophila)

TABLE S2

Let-7a/c predicted targets.

ADRB2	NM_000024	48	adrenergic, beta-2-,	hsa-miR-	−0.47	0.63	2005,
			receptor, surface	4458			2007,
							2009
ADRB3	NM_000025	49	adrenergic, beta-3-,	hsa-miR-	−0.28	0.98	2005,
			receptor	4458			2007,
							2009
FAS	NM_000043	50	Fas (TNF receptor	hsa-miR-98	−0.32	0.85	2009
			superfamily, member
			6)
ATP7B	NM_000053	51	ATPase, Cu++	hsa-miR-	−0.09	0.93	2009
			transporting, beta	4458
			polypeptide
CAPN3	NM_000070	52	calpain 3, (p94)	hsa-miR-	−0.15	0.92	2009
				4458
CLCN5	NM_000084	53	chloride channel 5	hsa-let-7c	−0.4	>0.99	2007,
							2009
COL1A1	NM_000088	54	collagen, type I, alpha 1	hsa-miR-	−0.18	0.89	2005,
				4500			2007,
							2009
COL1A2	NM_000089	55	collagen, type I, alpha 2	hsa-miR-	−0.41	0.95	2005,
				4500			2007,
							2009
COL3A1	NM_000090	56	collagen, type III,	hsa-miR-	−0.37	0.92	2005,
			alpha 1	4458			2007,
							2009
CYP19A1	NM_000103	57	cytochrome P450,	hsa-let-7f	−0.22	0.9	2005,
			family 19, subfamily				2007,
			A, polypeptide 1				2009
DMD	NM_000109	58	dystrophin	hsa-let-7d	−0.29	0.84	2005,
							2007,
							2009
ERCC6	NM_000124	59	excision repair cross-	hsa-let-7d	−0.46	0.95	2005,
			complementing				2007,
			rodent repair				2009
			deficiency,
			complementation
			group 6
GALC	NM_000153	60	galactosylceramidase	hsa-miR-	−0.27	0.93	2009
				4458
GHR	NM_000163	61	growth hormone	hsa-miR-98	−0.16	0.87	2005,
			receptor				2007,
							2009
HK2	NM_000189	62	hexokinase 2	hsa-miR-	−0.06	0.89	2005,
				4500			2007,
							2009
TBX5	NM_000192	63	T-box 5	hsa-miR-	−0.16	0.94	2005,
				4500			2007,
							2009
INSR	NM_000208	64	insulin receptor	hsa-let-7i	−0.11	0.99	2007,
							2009
ITGB3	NM_000212	65	integrin, beta 3	hsa-miR-	−0.21	>0.99	2005,
			(platelet glycoprotein	4458			2007,
			IIIa, antigen CD61)				2009
RB1	NM_000321	66	retinoblastoma 1	hsa-miR-	−0.1	0.83	2005,
				4500			2007,
							2009
SCN5A	NM_000335	67	sodium channel,	hsa-miR-	−0.04	0.95	2005,
			voltage-gated, type V,	4458			2007,
			alpha subunit				2009
SGCD	NM_000337	68	sarcoglycan, delta	hsa-miR-	>−0.02	0.7	2005,
			(35 kDa dystrophin-	4458			2007,
			associated				2009
			glycoprotein)
TSC1	NM_000368	69	tuberous sclerosis 1	hsa-miR-	−0.22	0.95	2005,
				4458			2007,
							2009
CDKN1A	NM_000389	70	cyclin-dependent	hsa-let-7i	−0.24	0.76	2009
			kinase inhibitor 1A
			(p21, Cip1)
TPP1	NM_000391	71	tripeptidyl peptidase I	hsa-miR-	−0.24	0.97	2009
				4458
COL5A2	NM_000393	72	collagen, type V,	hsa-miR-	−0.16	0.95	2005,
			alpha 2	4458			2007,
							2009
GALE	NM_000403	73	UDP-galactose-4-	hsa-miR-	−0.28	0.91	2005,
			epimerase	4458			2007,
							2009
LOR	NM_000427	74	loricrin	hsa-let-7g	−0.27	0.88	2009
RAG1	NM_000448	75	recombination	hsa-let-7a	−0.17	0.76	2009
			activating gene 1
AMT	NM_000481	76	aminomethyltransferase	hsa-let-7a	−0.29	0.82	2009
COL4A5	NM_000495	77	collagen, type IV,	hsa-miR-	−0.14	0.91	2005,
			alpha 5	4500			2007,
							2009
KCNJ11	NM_000525	78	potassium inwardly-	hsa-let-7d	−0.44	0.78	2009
			rectifying channel,
			subfamily J, member
			11
TP53	NM_000546	79	tumor protein p53	hsa-let-7d	−0.28	0.93	2009
DCX	NM_000555	80	doublecortin	hsa-let-7d	−0.05	0.87	2005,
							2007
IL6R	NM_000565	81	interleukin 6 receptor	hsa-miR-	−0.16	0.94
				4500
IL10	NM_000572	82	interleukin 10	hsa-let-7e	−0.14	0.94	2005,
							2007,
							2009
IL6	NM_000600	83	interleukin 6	hsa-miR-	−0.15	0.73	2007
			(interferon, beta 2)	4500
IGF1	NM_000618	84	insulin-like growth	hsa-let-7f	−0.1	0.97	2009
			factor 1
			(somatomedin C)
NOS1	NM_000620	85	nitric oxide synthase	hsa-miR-	−0.11	0.95
			1 (neuronal)	4458
FASLG	NM_000639	86	Fas ligand (TNF	hsa-miR-	−0.26	0.98	2005,
			superfamily, member	4458			2007,
			6)				2009
ADRB1	NM_000684	87	adrenergic, beta-1-,	hsa-miR-	−0.25	0.98	2007,
			receptor	4500			2009
CACNA1D	NM_000720	88	calcium channel,	hsa-let-7b	−0.07	0.88	2005,
			voltage-dependent, L				2007,
			type, alpha 1D				2009
			subunit
CACNA1E	NM_000721	89	calcium channel,	hsa-miR-	−0.01	0.93	2005,
			voltage-dependent, R	4500			2007,
			type, alpha 1E subunit				2009
GABRA6	NM_000811	90	gamma-aminobutyric	hsa-let-7d	−0.16	0.84	2009
			acid (GABA) A
			receptor, alpha 6
HTR1E	NM_000865	91	5-hydroxytryptamine	hsa-let-7d	−0.27	0.89	2009
			(serotonin) receptor
			1E
HTR4	NM_000870	92	5-hydroxytryptamine	hsa-let-7a	−0.12	0.94	2005,
			(serotonin) receptor 4				2007,
							2009
IGF1R	NM_000875	93	insulin-like growth	hsa-let-7b	−0.42	>0.99	2007,
			factor 1 receptor				2009
OPRM1	NM_000914	94	opioid receptor, mu 1	hsa-miR-	−0.2	0.92	2005,
				4500			2007,
							2009
PTAFR	NM_000952	95	platelet-activating	hsa-miR-	−0.69	0.94
			factor receptor	4500
NKIRAS2	NM_001001349	96	NFKB inhibitor	hsa-miR-	−0.09	0.88	2005,
			interacting Ras-like 2	4458			2007,
							2009
ATP2B4	NM_001001396	97	ATPase, Ca++	hsa-let-7f	−0.05	0.93	2005,
			transporting, plasma				2007,
			membrane 4				2009
RORC	NM_001001523	98	RAR-related orphan	hsa-miR-	−0.12	0.93	2005,
			receptor C	4458			2007,
							2009
GDF6	NM_001001557	99	growth differentiation	hsa-miR-	−0.5	0.98	2005,
			factor 6	4500			2007,
							2009
MTUS1	NM_001001924	100	microtubule	hsa-let-7d	−0.14	0.71	2009
			associated tumor
			suppressor 1
PPARA	NM_001001928	101	peroxisome	hsa-miR-	−0.04	0.97	2007,
			proliferator-activated	4458			2009
			receptor alpha
GOLGA7	NM_001002296	102	golgin A7	hsa-miR-	−0.15	0.7	2005,
				4500			2007,
							2009
WNK3	NM_001002838	103	WNK lysine deficient	hsa-miR-	−0.09	0.91
			protein kinase 3	4458
CACNA1I	NM_001003406	104	calcium channel,	hsa-miR-	−0.1	0.92	2009
			voltage-dependent, T	4458
			type, alpha 1I subunit
SMAD2	NM_001003652	105	SMAD family	hsa-miR-	−0.09	0.9	2007,
			member 2	4458			2009
C18orf1	NM_001003674	106	chromosome 18 open	hsa-miR-	−0.01	0.76
			reading frame 1	4500
KLHL31	NM_001003760	107	kelch-like 31	hsa-let-7d	−0.34	0.93	2009
			(Drosophila)
MGLL	NM_001003794	108	monoglyceride lipase	hsa-miR-	>−0.03	0.99	2005,
				4458			2007,
							2009
DLGAP1	NM_001003809	109	discs, large	hsa-miR-	−0.03	0.81
			(Drosophila)	4500
			homolog-associated
			protein 1
CD200	NM_001004196	110	CD200 molecule	hsa-miR-98	−0.12	0.75
ZNF740	NM_001004304	111	zinc finger protein	hsa-let-7d	>−0.02	0.94	2007,
			740				2009
LIN28B	NM_001004317	112	lin-28 homolog B (C. elegans)	hsa-let-7d	−0.98	>0.99	2007,
							2009
PLEKHG7	NM_001004330	113	pleckstrin homology	hsa-miR-	−0.34	0.94	2009
			domain containing,	4458
			family G (with
			RhoGef domain)
			member 7
TRIM67	NM_001004342	114	tripartite motif	hsa-let-7f	−0.08	>0.99	2007,
			containing 67				2009
NRARP	NM_001004354	115	NOTCH-regulated	hsa-miR-	−0.2	0.67	2005,
			ankyrin repeat protein	4458			2007,
							2009
ITGA11	NM_001004439	116	integrin, alpha 11	hsa-miR-	−0.07	0.77
				4458
YPEL2	NM_001005404	117	yippee-like 2	hsa-miR-	−0.15	0.96	2009
			(Drosophila)	4500
PLCXD3	NM_001005473	118	phosphatidylinositol-	hsa-miR-	−0.19	0.8	2005,
			specific	4458			2007
			phospholipase C, X
			domain containing 3
CPM	NM_001005502	119	carboxypeptidase M	hsa-miR-	−0.27	0.95	2005,
				4458			2007,
							2009
EDA	NM_001005609	120	ectodysplasin A	hsa-miR-	−0.06	0.92	2005,
				4458			2007,
							2009
ZNF473	NM_001006656	121	zinc finger protein	hsa-let-7d	−0.31	0.98	2009
			473
NTRK3	NM_001007156	122	neurotrophic tyrosine	hsa-let-7f	−0.16	0.82	2009
			kinase, receptor, type 3
IGF2BP2	NM_001007225	123	insulin-like growth	hsa-let-7b	−0.4	>0.99	2007,
			factor 2 mRNA				2009
			binding protein 2
BRWD1	NM_001007246	124	bromodomain and	hsa-let-7d	−0.5	0.73
			WD repeat domain
			containing 1
PRDM2	NM_001007257	125	PR domain containing	hsa-let-7g	−0.35	0.87
			2, with ZNF domain
GEMIN7	NM_001007269	126	gem (nuclear	hsa-miR-	−0.37	<0.1	2009
			organelle) associated	4500
			protein 7
IRGQ	NM_001007561	127	immunity-related	hsa-let-7g	−0.11	0.85
			GTPase family, Q
BCAP29	NM_001008405	128	B-cell receptor-	hsa-miR-	−0.16	0.94	2007,
			associated protein 29	4500			2009
STEAP3	NM_001008410	129	STEAP family	hsa-miR-	−0.28	0.98	2009
			member 3	4458
KIAA2022	NM_001008537	130	KIAA2022	hsa-let-7g	−0.12	0.92	2009
USP20	NM_001008563	131	ubiquitin specific	hsa-miR-	−0.06	0.84
			peptidase 20	4500
FNIP1	NM_001008738	132	folliculin interacting	hsa-miR-98	−0.22	0.98	2007,
			protein 1				2009
ACSL6	NM_001009185	133	acyl-CoA synthetase	hsa-let-7d	−0.39	0.98	2005,
			long-chain family				2007,
			member 6				2009
MFAP3L	NM_001009554	134	microfibrillar-	hsa-miR-	−0.09	0.93	2009
			associated protein 3-	4500
			like
MEIS3	NM_001009813	135	Meis homeobox 3	hsa-miR-	−0.26	0.92	2005,
				4458			2007,
							2009
KIAA0930	NM_001009880	136	KIAA0930	hsa-let-7d	−0.06	>0.99	2007,
							2009
C20orf194	NM_001009984	137	chromosome 20 open	hsa-miR-	−0.18	>0.99	2009
			reading frame 194	4500
ARHGAP28	NM_001010000	138	Rho GTPase	hsa-let-7a	−0.35	0.95	2005,
			activating protein 28				2007,
							2009
PM20D2	NM_001010853	139	peptidase M20	hsa-let-7a	−0.19	0.78
			domain containing 2
ACER2	NM_001010887	140	alkaline ceramidase 2	hsa-miR-	−0.15	0.94	2007,
				4458			2009
SLC5A9	NM_001011547	141	solute carrier family 5	hsa-miR-	−0.5	0.79	2009
			(sodium/glucose	4458
			cotransporter),
			member 9
NAA30	NM_001011713	142	N(alpha)-	hsa-miR-	−0.16	0.94	2007,
			acetyltransferase 30,	4500			2009
			NatC catalytic subunit
NHLRC3	NM_001012754	143	NHL repeat	hsa-let-7a	−0.27	0.95	2007,
			containing 3				2009
SNX30	NM_001012994	144	sorting nexin family	hsa-let-7d	−0.15	0.95	2009
			member 30
FIGNL2	NM_001013690	145	fidgetin-like 2	hsa-miR-	−1.07	>0.99	2009
				4458
C8orf58	NM_001013842	146	chromosome 8 open	hsa-let-7d	−0.42	0.97	2009
			reading frame 58
DCUN1D2	NM_001014283	147	DCN1, defective in	hsa-let-7b	−0.13	0.94	2007,
			cullin neddylation 1,				2009
			domain containing 2
			(S. cerevisiae)
KATNAL1	NM_001014380	148	katanin p60 subunit	hsa-miR-	−0.28	0.94	2009
			A-like 1	4458
USP21	NM_001014443	149	ubiquitin specific	hsa-miR-	−0.12	0.92	2005,
			peptidase 21	4458			2007,
							2009
DDX19B	NM_001014449	150	DEAD (Asp-Glu-Ala-	hsa-miR-	−0.47	0.97	2007,
			As) box polypeptide	4500			2009
			19B
RTKN	NM_001015055	151	rhotekin	hsa-miR-98	−0.14	0.76
GOPC	NM_001017408	152	golgi-associated PDZ	hsa-miR-	−0.1	0.87	2009
			and coiled-coil motif	4500
			containing
CALN1	NM_001017440	153	calneuron 1	hsa-miR-	−0.15	0.92	2009
				4458
C14orf28	NM_001017923	154	chromosome 14 open	hsa-let-7g	−1.25	>0.99	2007,
			reading frame 28				2009
OPA3	NM_001017989	155	optic atrophy 3	hsa-miR-	−0.42	0.71
			(autosomal recessive,	4458
			with chorea and
			spastic paraplegia)
DKK3	NM_001018057	156	dickkopf homolog 3	hsa-let-7d	−0.1	0.93	2007,
			(Xenopus laevis)				2009
SERF2	NM_001018108	157	small EDRK-rich	hsa-let-7d	−0.55	0.85
			factor 2
IQCB1	NM_001023570	158	IQ motif containing	hsa-miR-	−0.3	0.88	2009
			B1	4500
SBK1	NM_001024401	159	SH3-binding domain	hsa-let-7b	−0.1	0.94	2007,
			kinase 1				2009
CD276	NM_001024736	160	CD276 molecule	hsa-miR-	−0.06	0.71
				4458
BCL7A	NM_001024808	161	B-cell	hsa-miR-	−0.06	0.9	2005,
			CLL/lymphoma 7A	4500			2007,
							2009
KCTD21	NM_001029859	162	potassium channel	hsa-miR-98	−0.54	0.98	2007,
			tetramerisation				2009
			domain containing 21
SLC10A7	NM_001029998	163	solute carrier family	hsa-miR-98	−0.25	0.94
			10 (sodium/bile acid
			cotransporter family),
			member 7
CTNS	NM_001031681	164	cystinosin, lysosomal	hsa-let-7d	−0.12	0.93	2005,
			cystine transporter				2007,
							2009
RBFOX2	NM_001031695	165	RNA binding protein,	hsa-miR-98	−0.19	0.94	2005,
			fox-1 homolog (C. elegans) 2				2007,
							2009
PLD3	NM_001031696	166	phospholipase D	hsa-miR-	−0.13	0.9	2005,
			family, member 3	4458			2007,
							2009
ERGIC1	NM_001031711	167	endoplasmic	hsa-let-7d	−0.21	0.93
			reticulum-golgi
			intermediate
			compartment
			(ERGIC) 1
TMPO	NM_001032283	168	thymopoietin	hsa-miR-	−0.1	0.83
				4500
STK24	NM_001032296	169	serine/threonine	hsa-miR-	−0.11	0.93	2009
			kinase 24	4458
MYCL1	NM_001033081	170	v-myc	hsa-let-7a	−0.09	0.88	2007,
			myelocytomatosis				2009
			viral oncogene
			homolog 1, lung
			carcinoma derived
			(avian)
SRGAP3	NM_001033117	171	SLIT-ROBO Rho	hsa-miR-	−0.13	0.94	2005,
			GTPase activating	4500			2007,
			protein 3				2009
WIPI2	NM_001033518	172	WD repeat domain,	hsa-let-7e	−0.34	0.92	2009
			phosphoinositide
			interacting 2
RRM2	NM_001034	173	ribonucleotide	hsa-miR-	−0.36	0.95	2007,
			reductase M2	4500			2009
ARL5A	NM_001037174	174	ADP-ribosylation	hsa-let-7g	−0.13	0.94	2007,
			factor-like 5A				2009
CDC42SE1	NM_001038707	175	CDC42 small effector 1	hsa-miR-	−0.22	0.71	2009
				4500
TRIM71	NM_001039111	176	tripartite motif	hsa-miR-	−0.89	>0.99	2007,
			containing 71	4458			2009
TRIOBP	NM_001039141	177	TRIO and F-actin	hsa-miR-	−0.12	0.86	2009
			binding protein	4458
GK5	NM_001039547	178	glycerol kinase 5	hsa-let-7d	−0.16	0.85
			(putative)
KREMEN1	NM_001039570	179	kringle containing	hsa-miR-	−0.17	0.97	2007,
			transmembrane	4458			2009
			protein 1
SEC14L1	NM_001039573	180	SEC14-like 1 (S. cerevisiae)	hsa-miR-	−0.18	0.92	2005,
				4500			2007,
							2009
KCNC4	NM_001039574	181	potassium voltage-	hsa-miR-	−0.1	0.84	2009
			gated channel, Shaw-	4500
			related subfamily,
			member 4
TMPPE	NM_001039770	182	transmembrane	hsa-miR-	−0.17	0.84	2009
			protein with	4458
			metallophosphoesterase
			domain
CHIC1	NM_001039840	183	cysteine-rich	hsa-miR-	>−0.04	>0.99	2009
			hydrophobic domain 1	4458
MLLT4	NM_001040000	184	myeloid/lymphoid or	hsa-miR-	−0.16	0.74	2007,
			mixed-lineage	4458			2009
			leukemia (trithorax
			homolog,
			Drosophila);
			translocated to, 4
PQLC2	NM_001040125	185	PQ loop repeat	hsa-miR-	−0.3	0.95	2009
			containing 2	4458
PEG10	NM_001040152	186	paternally expressed	hsa-miR-	−0.02	0.71	2009
			10	4458
MTMR12	NM_001040446	187	myotubularin related	hsa-miR-	−0.06	0.9	2009
			protein 12	4458
PTPRD	NM_001040712	188	protein tyrosine	hsa-miR-	−0.22	0.96	2007,
			phosphatase, receptor	4500			2009
			type, D
KIAA0895L	NM_001040715	189	KIAA0895-like	hsa-let-7i	−0.14	0.93	2007,
							2009
USP44	NM_001042403	190	ubiquitin specific	hsa-miR-	−0.14	0.93	2009
			peptidase 44	4458
RUFY2	NM_001042417	191	RUN and FYVE	hsa-miR-98	−0.27	0.82
			domain containing 2
C6orf204	NM_001042475	192	chromosome 6 open	hsa-miR-	−0.13	0.94
			reading frame 204	4458
DLGAP4	NM_001042486	193	discs, large	hsa-let-7d	−0.31	0.97	2009
			(Drosophila)
			homolog-associated
			protein 4
C2orf88	NM_001042519	194	chromosome 2 open	hsa-miR-	−0.17	0.89	2009
			reading frame 88	4500
ATPAF1	NM_001042546	195	ATP synthase	hsa-let-7d	−0.43	0.92	2009
			mitochondrial F1
			complex assembly
			factor 1
FRS2	NM_001042555	196	fibroblast growth	hsa-let-7i	−0.04	0.86	2007,
			factor receptor				2009
			substrate 2
EIF4G2	NM_001042559	197	eukaryotic translation	hsa-let-7d	−0.27	0.92	2005,
			initiation factor 4				2007,
			gamma, 2				2009
DPH3	NM_001047434	198	DPH3, KTI11	hsa-let-7i	−0.4	0.98
			homolog (S. cerevisiae)
UHRF1	NM_001048201	199	ubiquitin-like with	hsa-let-7d	−0.17	0.94	2009
			PHD and ring finger
			domains 1
TNFRSF1B	NM_001066	200	tumor necrosis factor	hsa-miR-	−0.09	0.99	2005,
			receptor superfamily,	4458			2007,
			member 1B				2009
CDC14B	NM_001077181	201	CDC14 cell division	hsa-miR-	−0.08	0.88	2009
			cycle 14 homolog B	4500
			(S. cerevisiae)
ATG9A	NM_001077198	202	ATG9 autophagy	hsa-let-7d	−0.03	0.74
			related 9 homolog A
			(S. cerevisiae)
SREK1	NM_001077199	203	splicing regulatory	hsa-miR-	−0.09	0.92	2005,
			glutamine/lysine-rich	4458			2007,
			protein 1				2009
IKZF2	NM_001079526	204	IKAROS family zinc	hsa-let-7g	−0.13	0.98	2007,
			finger 2 (Helios)				2009
CPEB1	NM_001079533	205	cytoplasmic	hsa-miR-	−0.44	0.91	2007,
			polyadenylation	4500			2009
			element binding
			protein 1
FNDC3A	NM_001079673	206	fibronectin type III	hsa-let-7d	−0.39	0.97	2005,
			domain containing 3A				2007,
							2009
GYG2	NM_001079855	207	glycogenin 2	hsa-let-7a	−0.22	0.88	2009
VAV3	NM_001079874	208	vav 3 guanine	hsa-miR-	−0.11	0.92	2005,
			nucleotide exchange	4500			2007,
			factor				2009
DMP1	NM_001079911	209	dentin matrix acidic	hsa-miR-	−0.21	0.94	2005,
			phosphoprotein 1	4500			2007,
							2009
LDB3	NM_001080114	210	LIM domain binding 3	hsa-miR-	−0.09	0.84	2009
				4458
GJC1	NM_001080383	211	gap junction protein,	hsa-miR-	−0.42	0.95	2009
			gamma 1, 45 kDa	4500
KIAA1147	NM_001080392	212	KIAA1147	hsa-miR-	−0.15	0.86	2007,
				4458			2009
SLC45A4	NM_001080431	213	solute carrier family	hsa-let-7d	−0.08	0.91	2003,
			45, member 4				2007,
							2009
DNA2	NM_001080449	214	DNA replication	hsa-miR-	−0.53	0.6	2009
			helicase 2 homolog	4458
			(yeast)
MYO5B	NM_001080467	215	myosin VB	hsa-let-7g	−0.04	0.78	2009
ZNF697	NM_001080470	216	zinc finger protein	hsa-miR-	−0.14	0.93	2007,
			697	4458			2009
ZNF275	NM_001080485	217	zinc finger protein	hsa-miR-	−0.11	0.99	2007,
			275	4458			2009
MGA	NM_001080541	218	MAX gene associated	hsa-miR-	−0.12	0.91	2007,
				4500			2009
THOC2	NM_001081550	219	THO complex 2	hsa-miR-	−0.17	0.85	2007,
				4458			2009
CPSF4	NM_001081559	220	cleavage and	hsa-miR-	−0.12	0.77	2005,
			polyadenylation	4458			2007,
			specific factor 4,				2009
			30 kDa
C11orf57	NM_001082969	221	chromosome 11 open	hsa-let-7f	−0.25	0.89	2007,
			reading frame 57				2009
E2F5	NM_001083588	222	E2F transcription	hsa-miR-	−0.3	0.86	2009
			factor 5, p130-binding	4500
ANKRD12	NM_001083625	223	ankyrin repeat	hsa-miR-	>−0.02	0.77
			domain 12	4458
FOXN3	NM_001085471	224	forkhead box N3	hsa-miR-	>−0.01	0.82
				4458
MEX3A	NM_001093725	225	mex-3 homolog A (C. elegans)	hsa-miR-98	−0.15	0.92	2009
HIC1	NM_001098202	226	hypermethylated in	hsa-miR-	−0.05	0.84	2009
			cancer 1	4458
MGAT3	NM_001098270	227	mannosyl (beta-1,4-)-	hsa-miR-	>−0.02	0.66
			glycoprotein beta-1,4-	4458
			N-
			acetylglucosaminyltransferase
SLC4A4	NM_001098484	228	solute carrier family	hsa-miR-	−0.16	0.95	2005,
			4, sodium bicarbonate	4458			2007,
			cotransporter,				2009
			member 4
FAM104A	NM_001098832	229	family with sequence	hsa-miR-	−0.21	0.92	2007,
			similarity 104,	4458			2009
			member A
ATXN7L3	NM_001098833	230	ataxin 7-like 3	hsa-miR-	−0.06	0.74
				4500
BTBD9	NM_001099272	231	BTB (POZ) domain	hsa-let-7a	−0.16	0.95	2009
			containing 9
NIPAL4	NM_001099287	232	NIPA-like domain	hsa-miR-	−0.17	0.93	2009
			containing 4	4458
SH3RF3	NM_001099289	233	SH3 domain	hsa-miR-	−0.08	0.92	2009
			containing ring finger 3	4458
GXYLT1	NM_001099650	234	glucoside	hsa-miR-	−0.37	0.99	2009
			xylosyltransferase 1	4500
PTAR1	NM_001099666	235	protein	hsa-miR-	−0.15	0.98	2009
			prenyltransferase	4500
			alpha subunit repeat
			containing 1
FBXL19	NM_001099784	236	F-box and leucine-	hsa-miR-	−0.1	0.7	2009
			rich repeat protein 19	4458
ACTA1	NM_001100	237	actin, alpha 1, skeletal	hsa-let-7c	−0.21	0.72	2005,
			muscle				2007
PHACTR2	NM_001100164	238	phosphatase and actin	hsa-miR-	−0.15	0.94	2009
			regulator 2	4500
MOBKL3	NM_001100819	239	MOB1, Mps One	hsa-let-7a	−0.12	0.93	2005,
			Binder kinase				2007,
			activator-like 3				2009
			(yeast)
PACS2	NM_001100913	240	phosphofurin acidic	hsa-miR-	−0.14	0.79	2009
			cluster sorting protein 2	4458
IGLON5	NM_001101372	241	IgLON family	hsa-let-7d	−0.04	0.94	2009
			member 5
SAMD12	NM_001101676	242	sterile alpha motif	hsa-miR-	−0.05	0.94	2009
			domain containing 12	4500
DTX2	NM_001102594	243	deltex homolog 2	hsa-let-7d	−0.46	>0.99	2005,
			(Drosophila)				2007,
							2009
FAM118A	NM_001104595	244	family with sequence	hsa-miR-	−0.28	0.98	2007,
			similarity 118,	4500			2009
			member A
FAM123C	NM_001105193	245	family with sequence	hsa-miR-	−0.25	0.87	2009
			similarity 123C	4500
PCDH19	NM_001105243	246	protocadherin 19	hsa-miR-	−0.12	0.95	2007,
				4500			2009
ZFYVE16	NM_001105251	247	zinc finger, FYVE	hsa-miR-	−0.16	0.71	2009
			domain containing 16	4500
SWT1	NM_001105518	248	SWT1 RNA	hsa-let-7b	−0.21	0.85	2007,
			endoribonuclease				2009
			homolog (S. cerevisiae)
CAP1	NM_001105530	249	CAP, adenylate	hsa-miR-	−0.14	0.85	2005,
			cyclase-associated	4500			2007,
			protein 1 (yeast)				2009
FAM135A	NM_001105531	250	family with sequence	hsa-let-7f	−0.19	0.85	2005,
			similarity 135,				2007,
			member A				2009
ZBTB10	NM_001105539	251	zinc finger and BTB	hsa-miR-	−0.06	0.72	2005,
			domain containing 10	4500			2007
NEFM	NM_001105541	252	neurofilament,	hsa-let-7f	−0.16	0.86	2007,
			medium polypeptide				2009
FBXO45	NM_001105573	253	F-box protein 45	hsa-miR-	−0.06	0.89	2009
				4458
ACVR2B	NM_001106	254	activin A receptor,	hsa-miR-	−0.03	>0.99	2007,
			type IIB	4500			2009
C12orf51	NM_001109662	255	chromosome 12 open	hsa-miR-	−0.09	0.92	2009
			reading frame 51	4458
GSG1L	NM_001109763	256	GSG1-like	hsa-miR-	−0.08	0.73
				4458
ACVR1C	NM_001111031	257	activin A receptor,	hsa-miR-	−0.48	0.99	2005,
			type IC	4458			2007,
							2009
C3orf63	NM_001112736	258	chromosome 3 open	hsa-miR-	−0.02	0.91	2009
			reading frame 63	4458
HIPK2	NM_001113239	259	homeodomain	hsa-let-7d	−0.03	0.94	2009
			interacting protein
			kinase 2
AMOT	NM_001113490	260	angiomotin	hsa-miR-	−0.12	0.76
				4458
ARHGEF7	NM_001113513	261	Rho guanine	hsa-let-7a	−0.14	0.94
			nucleotide exchange
			factor (GEF) 7
CTSC	NM_001114173	262	cathepsin C	hsa-miR-	−0.15	0.89	2009
				4458
ADCY9	NM_001116	263	adenylate cyclase 9	hsa-miR-	−0.03	0.91	2005,
				4458			2007,
							2009
NAPEPLD	NM_001122838	264	N-acyl	hsa-miR-	−0.05	0.94	2007,
			phosphatidylethanolamine	4458			2009
			phospholipase D
CASK	NM_001126054	265	calcium/calmodulin-	hsa-let-7d	−0.04	0.91
			dependent serine
			protein kinase
			(MAGUK family)
ELF4	NM_001127197	266	E74-like factor 4 (ets	hsa-let-7a	−0.17	0.94	2007,
			domain transcription				2009
			factor)
TET2	NM_001127208	267	tet oncogene family	hsa-miR-	−0.16	0.96
			member 2	4458
CBX5	NM_001127321	268	chromobox homolog 5	hsa-miR-	−0.17	>0.99
				4458
CRY2	NM_001127457	269	cryptochrome 2	hsa-miR-	−0.12	0.93	2005,
			(photolyase-like)	4458			2007,
							2009
STXBP5	NM_001127715	270	syntaxin binding	hsa-let-7d	−0.15	0.8	2009
			protein 5 (tomosyn)
SULF1	NM_001128204	271	sulfatase 1	hsa-miR-	−0.1	0.85	2009
				4500
SMARCAD1	NM_001128429	272	SWI/SNF-related,	hsa-let-7f	−0.57	0.95	2005,
			matrix-associated				2007,
			actin-dependent				2009
			regulator of
			chromatin, subfamily
			a, containing
			DEAD/H box 1
RASGRP1	NM_001128602	273	RAS guanyl releasing	hsa-miR-	−0.34	0.97	2005,
			protein 1 (calcium	4458			2007,
			and DAG-regulated)				2009
PAK1	NM_001128620	274	p21 protein	hsa-miR-	−0.18	0.78	2005,
			(Cdc42/Rac)-	4500			2007,
			activated kinase 1				2009
SPIRE1	NM_001128626	275	spire homolog 1	hsa-let-7a	−0.04	0.92	2009
			(Drosophila)
CALU	NM_001130674	276	calumenin	hsa-miR-	−0.19	0.93	2005,
				4458			2007,
							2009
STYX	NM_001130701	277	serine/threonine/tyrosine	hsa-miR-	−0.09	0.93
			interacting protein	4500
GAS7	NM_001130831	278	growth arrest-specific 7	hsa-miR-	−0.28	0.98	2005,
				4500			2007,
							2009
RTCD1	NM_001130841	279	RNA terminal	hsa-miR-	−0.2	0.88
			phosphate cyclase	4458
			domain 1
TGFBR1	NM_001130916	280	transforming growth	hsa-let-7f	−0.52	>0.99	2005,
			factor, beta receptor 1				2007,
							2009
TTLL6	NM_001130918	281	tubulin tyrosine	hsa-miR-	−0.26	0.76
			ligase-like family,	4458
			member 6
TMEM194A	NM_001130963	282	transmembrane	hsa-let-7a	−0.07	0.94	2009
			protein 194A
MEF2C	NM_001131005	283	myocyte enhancer	hsa-miR-	−0.24	0.87
			factor 2C	4458
SAP30L	NM_001131062	284	SAP30-like	hsa-miR-	>−0.02	0.92
				4458
CCNJ	NM_001134375	285	cyclin J	hsa-miR-	−0.44	0.94	2005,
				4458			2007,
							2009
CYB561D1	NM_001134400	286	cytochrome b-561	hsa-let-7d	−0.33	0.91
			domain containing 1
CDV3	NM_001134422	287	CDV3 homolog	hsa-miR-	−0.13	0.98	2007,
			(mouse)	4500			2009
LRRC8B	NM_001134476	288	leucine rich repeat	hsa-miR-	−0.07	0.88
			containing 8 family,	4500
			member B
PBX3	NM_001134778	289	pre-B-cell leukemia	hsa-miR-98	−0.32	0.99	2005,
			homeobox 3				2007,
							2009
FNDC3B	NM_001135095	290	fibronectin type III	hsa-miR-	−0.16	0.98	2005,
			domain containing 3B	4500			2007,
							2009
TMPRSS2	NM_001135099	291	transmembrane	hsa-let-7b	−0.32	0.98	2007,
			protease, serine 2				2009
HDHD1	NM_001135565	292	haloacid	hsa-miR-	−0.28	0.62	2005,
			dehalogenase-like	4500			2007,
			hydrolase domain				2009
			containing 1
LOC221710	NM_001135575	293	hypothetical protein	hsa-miR-	−0.01	0.78
			LOC221710	4458
SPATA2	NM_001135773	294	spermatogenesis	hsa-let-7a	−0.07	0.94	2005,
			associated 2				2007,
							2009
C9orf7	NM_001135775	295	chromosome 9 open	hsa-miR-	−0.12	0.92	2005,
			reading frame 7	4500			2007,
							2009
SYT1	NM_001135805	296	synaptotagmin I	hsa-miR-	−0.16	0.87	2005,
				4458			2007,
							2009
TMEM2	NM_001135820	297	transmembrane	hsa-let-7g	−0.41	0.98	2005,
			protein 2				2007,
							2009
PIK3IP1	NM_001135911	298	phosphoinositide-3-	hsa-let-7a	−0.19	0.85	2005,
			kinase interacting				2007,
			protein 1				2009
TTC39C	NM_001135993	299	tetratricopeptide	hsa-miR-	−0.06	0.86
			repeat domain 39C	4500
ABL2	NM_001136000	300	v-abl Abelson murine	hsa-let-7f	−0.23	>0.99	2009
			leukemia viral
			oncogene homolog 2
MICAL3	NM_001136004	301	microtubule	hsa-let-7g	−0.22	0.83
			associated
			monoxygenase,
			calponin and LIM
			domain containing 3
LMLN	NM_001136049	302	leishmanolysin-like	hsa-miR-	−0.13	0.9
			(metallopeptidase M8	4500
			family)
LRIG3	NM_001136051	303	leucine-rich repeats	hsa-miR-	−0.49	0.96	2005,
			and immunoglobulin-	4458			2007,
			like domains 3				2009
ZNF879	NM_001136116	304	zinc finger protein	hsa-miR-	−0.34	0.95
			879	4458
ATXN7L3B	NM_001136262	305	ataxin 7-like 3B	hsa-miR-	>−0.01	0.91
				4458
VASH2	NM_001136474	306	vasohibin 2	hsa-let-7d	−0.13	0.93	2007,
							2009
BTF3L4	NM_001136497	307	basic transcription	hsa-miR-	−0.12	0.9	2009
			factor 3-like 4	4458
SYT2	NM_001136504	308	synaptotagmin II	hsa-miR-	−0.16	0.94	2009
				4500
ATXN1L	NM_001137675	309	ataxin 1-like	hsa-miR-	>−0.02	0.93
				4458
NIPA1	NM_001142275	310	non imprinted in	hsa-miR-	−0.2	0.99	2007,
			Prader-	4458			2009
			Willi/Angelman
			syndrome 1
RBFOX1	NM_001142333	311	RNA binding protein,	hsa-let-7f	−0.18	0.86	2005,
			fox-1 homolog (C. elegans) 1				2007,
							2009
GNAL	NM_001142339	312	guanine nucleotide	hsa-miR-	−0.13	0.95	2005,
			binding protein (G	4500			2007,
			protein), alpha				2009
			activating activity
			polypeptide, olfactory
			type
SCN4B	NM_001142348	313	sodium channel,	hsa-miR-98	−0.08	0.93	2009
			voltage-gated, type
			IV, beta
CTIF	NM_001142397	314	CBP80/20-dependent	hsa-let-7a	−0.1	0.77
			translation initiation
			factor
RPUSD3	NM_001142547	315	RNA pseudouridylate	hsa-miR-	−0.42	0.89	2007,
			synthase domain	4500			2009
			containing 3
BBX	NM_001142568	316	bobby sox homolog	hsa-let-7d	−0.11	0.81
			(Drosophila)
CLP1	NM_001142597	317	CLP1, cleavage and	hsa-miR-	−0.32	0.68	2007
			polyadenylation	4458
			factor I subunit,
			homolog (S. cerevisiae)
TMOD2	NM_001142885	318	tropomodulin 2	hsa-miR-	−0.12	0.98
			(neuronal)	4458
PLEKHG6	NM_001144856	319	pleckstrin homology	hsa-miR-	−0.37	0.98	2007,
			domain containing,	4458			2009
			family G (with
			RhoGef domain)
			member 6
LIPT2	NM_001144869	320	lipoyl(octanoyl)	hsa-miR-	−0.43	0.87
			transferase 2	4458
			(putative)
SLC30A7	NM_001144884	321	solute carrier family	hsa-let-7d	−0.07	0.9	2009
			30 (zinc transporter),
			member 7
NEDD4L	NM_001144964	322	neural precursor cell	hsa-miR-	−0.06	0.94
			expressed,	4500
			developmentally
			down-regulated 4-like
PALM3	NM_001145028	323	paralemmin 3	hsa-miR-	−0.12	0.91
				4458
LRRC10B	NM_001145077	324	leucine rich repeat	hsa-let-7a	−0.07	0.62
			containing 10B
GRID2IP	NM_001145118	325	glutamate receptor,	hsa-miR-	−0.26	0.94
			ionotropic, delta 2	4458
			(Grid2) interacting
			protein
CDK6	NM_001145306	326	cyclin-dependent	hsa-miR-	−0.08	0.85
			kinase 6	4458
ZNF566	NM_001145343	327	zinc finger protein	hsa-let-7f	−0.25	0.93	2009
			566
ZNF652	NM_001145365	328	zinc finger protein	hsa-miR-	−0.15	>0.99
			652	4458
POTEM	NM_001145442	329	POTE ankyrin	hsa-miR-	−0.22	<0.1
			domain family,	4500
			member M
ZNF200	NM_001145446	330	zinc finger protein	hsa-let-7d	−0.37	>0.99	2009
			200
SOX6	NM_001145811	331	SRY (sex determining	hsa-let-7d	−0.16	0.72
			region Y)-box 6
SLCO5A1	NM_001146008	332	solute carrier organic	hsa-miR-	−0.08	0.81	2005,
			anion transporter	4500			2007,
			family, member 5A1				2009
NEK3	NM_001146099	333	NIMA (never in	hsa-miR-	−0.35	0.91	2009
			mitosis gene a)-	4500
			related kinase 3
SLC6A15	NM_001146335	334	solute carrier family 6	hsa-miR-	−0.15	0.94
			(neutral amino acid	4500
			transporter), member
			15
SLC25A4	NM_001151	335	solute carrier family	hsa-miR-	−0.15	0.94	2005,
			25 (mitochondrial	4500			2007
			carrier; adenine
			nucleotide
			translocator), member 4
KLF8	NM_001159296	336	Kruppel-like factor 8	hsa-miR-	−0.11	0.81
				4458
BEGAIN	NM_001159531	337	brain-enriched	hsa-miR-	−0.36	0.99	2005,
			guanylate kinase-	4458			2007,
			associated homolog				2009
			(rat)
BEND4	NM_001159547	338	BEN domain	hsa-miR-98	−0.24	>0.99	2009
			containing 4
SYNCRIP	NM_001159673	339	synaptotagmin	hsa-miR-	−0.29	0.96
			binding, cytoplasmic	4458
			RNA interacting
			protein
BZW2	NM_001159767	340	basic leucine zipper	hsa-let-7a	−0.24	0.81	2007,
			and W2 domains 2				2009
CANT1	NM_001159772	341	calcium activated	hsa-miR-	−0.19	0.89	2009
			nucleotidase 1	4458
ZNF583	NM_001159860	342	zinc finger protein	hsa-let-7f	−0.44	0.98	2007,
			583				2009
RNF170	NM_001160223	343	ring finger protein	hsa-let-7d	−0.41	0.98
			170
PANX2	NM_001160300	344	pannexin 2	hsa-miR-	−0.1	0.92	2005,
				4458			2007,
							2009
TMC7	NM_001160364	345	transmembrane	hsa-let-7d	−0.12	0.92	2005,
			channel-like 7				2007,
							2009
IGF2BP1	NM_001160423	346	insulin-like growth	hsa-miR-	−0.4	>0.99	2007,
			factor 2 mRNA	4500			2009
			binding protein 1
SYNC	NM_001161708	347	syncoilin,	hsa-miR-	−0.48	<0.1
			intermediate filament	4500
			protein
SULF2	NM_001161841	348	sulfatase 2	hsa-miR-	−0.12	0.91
				4458
APBA1	NM_001163	349	amyloid beta (A4)	hsa-let-7a	−0.11	0.94
			precursor protein-
			binding, family A,
			member 1
CECR6	NM_001163079	350	cat eye syndrome	hsa-let-7d	−0.14	0.95	2005,
			chromosome region,				2007,
			candidate 6				2009
PCYT1B	NM_001163264	351	phosphate	hsa-miR-	>−0.02	0.91	2007,
			cytidylyltransferase 1,	4458			2009
			choline, beta
CPA4	NM_001163446	352	carboxypeptidase A4	hsa-miR-	−0.37	0.93	2005,
				4458			2007,
							2009
GTF2I	NM_001163636	353	general transcription	hsa-miR-	−0.2	0.85	2007,
			factor IIi	4458			2009
DLC1	NM_001164271	354	deleted in liver cancer 1	hsa-miR-	−0.22	>0.99	2005,
				4458			2007,
							2009
SLC37A4	NM_001164277	355	solute carrier family	hsa-let-7d	−0.11	0.7
			37 (glucose-6-
			phosphate
			transporter), member 4
ANKRD33B	NM_001164440	356	ankyrin repeat	hsa-miR-	>−0.03	0.25
			domain 33B	4458
C16orf52	NM_001164579	357	chromosome 16 open	hsa-miR-	−0.09	0.83
			reading frame 52	4458
KIAA1549	NM_001164665	358	KIAA1549	hsa-miR-	>−0.02	0.94	2009
				4458
PITPNM3	NM_001165966	359	PITPNM family	hsa-miR-	>−0.02	0.94
			member 3	4458
EPB41	NM_001166005	360	erythrocyte	hsa-let-7a	−0.06	0.76	2009
			membrane protein
			band 4.1
			(elliptocytosis 1, RH-
			linked)
CEP120	NM_001166226	361	centrosomal protein	hsa-miR-	−0.22	0.92	2007,
			120 kDa	4458			2009
GRIK2	NM_001166247	362	glutamate receptor,	hsa-miR-	−0.12	0.86	2005,
			ionotropic, kainate 2	4500			2007,
							2009
SPATA13	NM_001166271	363	spermatogenesis	hsa-miR-	>−0.02	0.64
			associated 13	4458
TRAPPC1	NM_001166621	364	trafficking protein	hsa-miR-	N/A	0.12	2005,
			particle complex 1	4458			2007,
							2009
TMED5	NM_001167830	365	transmembrane	hsa-miR-	−0.11	0.95	2005,
			emp24 protein	4500			2007,
			transport domain				2009
			containing 5
SBNO1	NM_001167856	366	strawberry notch	hsa-miR-	−0.17	0.95
			homolog 1	4458
			(Drosophila)
TIMM17B	NM_001167947	367	translocase of inner	hsa-miR-	−0.16	0.93	2005,
			mitochondrial	4458			2007,
			membrane 17				2009
			homolog B (yeast)
GPR156	NM_001168271	368	G protein-coupled	hsa-miR-	−0.21	0.95
			receptor 156	4458
EDN1	NM_001168319	369	endothelin 1	hsa-miR-	−0.31	0.86	2009
				4500
TXLNG	NM_001168683	370	taxilin gamma	hsa-miR-	−0.32	0.95
				4500
TMEM135	NM_001168724	371	transmembrane	hsa-let-7a	−0.11	0.93
			protein 135
AFF2	NM_001169122	372	AF4/FMR2 family,	hsa-miR-	−0.22	0.92	2009
			member 2	4458
GPR137	NM_001170726	373	G protein-coupled	hsa-let-7b	−0.16	0.67	2003,
			receptor 137				2007,
							2009
LCOR	NM_001170765	374	ligand dependent	hsa-miR-	−0.14	0.96	2007,
			nuclear receptor	4500			2009
			corepressor
IGSF1	NM_001170963	375	immunoglobulin	hsa-let-7f	−0.38	0.93	2009
			superfamily, member 1
STRBP	NM_001171137	376	spermatid perinuclear	hsa-miR-	−0.2	0.89	2005,
			RNA binding protein	4458			2007,
							2009
C3orf52	NM_001171747	377	chromosome 3 open	hsa-miR-	−0.23	0.85	2009
			reading frame 52	4458
CSRNP3	NM_001172173	378	cysteine-serine-rich	hsa-let-7c	−0.05	0.87
			nuclear protein 3
OLR1	NM_001172632	379	oxidized low density	hsa-miR-	−0.12	0.88	2005,
			lipoprotein (lectin-	4458			2007,
			like) receptor 1				2009
RAB40C	NM_001172663	380	RAB40C, member	hsa-let-7a	−0.1	0.91	2005,
			RAS oncogene family				2007,
							2009
ZNF347	NM_001172674	381	zinc finger protein	hsa-miR-	−0.41	<0.1
			347	4458
ZNF641	NM_001172681	382	zinc finger protein	hsa-miR-	−0.2	0.93
			641	4458
PPARGC1B	NM_001172698	383	peroxisome	hsa-miR-	−0.31	>0.99	2005,
			proliferator-activated	4500			2007,
			receptor gamma,				2009
			coactivator 1 beta
PPP1R16B	NM_001172735	384	protein phosphatase 1,	hsa-miR-	−0.07	0.94	2005,
			regulatory (inhibitor)	4458			2007,
			subunit 16B				2009
FOXP2	NM_001172766	385	forkhead box P2	hsa-miR-	−0.33	>0.99
				4500
CRBN	NM_001173482	386	cereblon	hsa-miR-	−0.2	0.74
				4458
LMX1A	NM_001174069	387	LIM homeobox	hsa-miR-98	−0.16	0.92	2007,
			transcription factor 1,				2009
			alpha
POLL	NM_001174084	388	polymerase (DNA	hsa-miR-	−0.31	<0.1	2009
			directed), lambda	4458
NCOA3	NM_001174087	389	nuclear receptor	hsa-miR-	−0.1	0.84	2005,
			coactivator 3	4458			2007
TRPM6	NM_001177310	390	transient receptor	hsa-miR-	−0.2	0.98	2005,
			potential cation	4458			2007,
			channel, subfamily				2009
			M, member 6
CPEB2	NM_001177381	391	cytoplasmic	hsa-let-7b	−0.21	0.98	2005,
			polyadenylation				2007,
			element binding				2009
			protein 2
HDX	NM_001177478	392	highly divergent	hsa-let-7g	−0.38	0.97
			homeobox
PPP2R2A	NM_001177591	393	protein phosphatase 2,	hsa-miR-	−0.19	0.94
			regulatory subunit B,	4458
			alpha
STX3	NM_001178040	394	syntaxin 3	hsa-let-7g	−0.41	0.98	2007,
							2009
PARP8	NM_001178055	395	poly (ADP-ribose)	hsa-miR-	−0.23	0.98
			polymerase family,	4458
			member 8
BCAT1	NM_001178091	396	branched chain	hsa-miR-	−0.1	0.95	2009
			amino-acid	4500
			transaminase 1,
			cytosolic
CPEB3	NM_001178137	397	cytoplasmic	hsa-miR-	−0.15	0.96	2005,
			polyadenylation	4458			2007,
			element binding				2009
			protein 3
SLAMF6	NM_001184714	398	SLAM family	hsa-miR-	−0.22	0.88	2007,
			member 6	4458			2009
PEX11B	NM_001184795	399	peroxisomal	hsa-miR-	−0.34	0.51	2009
			biogenesis factor 11	4458
			beta
PHF8	NM_001184896	400	PHD finger protein 8	hsa-miR-	>−0.02	0.92	2005,
				4458			2007,
							2009
CLDN12	NM_001185072	401	claudin 12	hsa-miR-	−0.34	0.98	2005,
				4458			2007,
							2009
BACH1	NM_001186	402	BTB and CNC	hsa-let-7c	−0.31	>0.99	2005,
			homology 1, basic				2007,
			leucine zipper				2009
			transcription factor 1
ATG16L1	NM_001190266	403	ATG16 autophagy	hsa-miR-	−0.09	0.92	2007,
			related 16-like 1 (S. cerevisiae)	4458			2009
NCOR1	NM_001190440	404	nuclear receptor	hsa-miR-	−0.05	0.92
			corepressor 1	4458
COL11A1	NM_001190709	405	collagen, type XI,	hsa-miR-	−0.1	0.77
			alpha 1	4458
YAF2	NM_001190977	406	YY1 associated factor 2	hsa-let-7a	−0.05	0.94	2009
BCL2L1	NM_001191	407	BCL2-like 1	hsa-miR-	−0.2	0.9	2005,
				4458			2007,
							2009
IKBKE	NM_001193321	408	inhibitor of kappa	hsa-let-7b	−0.13	0.94	2005,
			light polypeptide gene				2007,
			enhancer in B-cells,				2009
			kinase epsilon
SECISBP2L	NM_001193489	409	SECIS binding	hsa-miR-	−0.08	0.81
			protein 2-like	4458
SLC1A4	NM_001193493	410	solute carrier family 1	hsa-let-7d	−0.05	0.98	2009
			(glutamate/neutral
			amino acid
			transporter), member 4
SLC30A6	NM_001193513	411	solute carrier family	hsa-miR-	−0.31	0.92
			30 (zinc transporter),	4458
			member 6
POGZ	NM_001194937	412	pogo transposable	hsa-miR-	−0.16	0.88	2005,
			element with ZNF	4458			2007,
			domain				2009
PTPRU	NM_001195001	413	protein tyrosine	hsa-let-7a	−0.14	0.92	2009
			phosphatase, receptor
			type, U
ANKRD28	NM_001195098	414	ankyrin repeat	hsa-miR-	−0.2	0.84	2009
			domain 28	4458
PTP4A2	NM_001195100	415	protein tyrosine	hsa-let-7b	−0.07	0.75
			phosphatase type
			IVA, member 2
LOC100507421	NM_001195278	416	transmembrane	hsa-miR-	−0.06	0.85
			protein 178-like	4458
DICER1	NM_001195573	417	dicer 1, ribonuclease	hsa-miR-	−0.05	>0.99	2009
			type III	4458
MLLT10	NM_001195626	418	myeloid/lymphoid or	hsa-let-7i	−0.15	0.86	2005,
			mixed-lineage				2007,
			leukemia (trithorax				2009
			homolog,
			Drosophila);
			translocated to, 10
TGFBR3	NM_001195683	419	transforming growth	hsa-let-7g	−0.39	0.98
			factor, beta receptor
			III
PLD5	NM_001195811	420	phospholipase D	hsa-miR-98	−0.18	0.86
			family, member 5
PLEKHA8	NM_001197026	421	pleckstrin homology	hsa-let-7a	−0.32	0.99
			domain containing,
			family A
			(phosphoinositide
			binding specific)
			member 8
DPYSL3	NM_001197294	422	dihydropyrimidinase-	hsa-miR-	−0.03	0.84	2009
			like 3	4500
GABPA	NM_001197297	423	GA binding protein	hsa-miR-	−0.09	0.92	2007,
			transcription factor,	4500			2009
			alpha subunit 60 kDa
PRDM1	NM_001198	424	PR domain containing	hsa-let-7a	−0.05	0.74	2005,
			1, with ZNF domain				2007,
							2009
RUNX1T1	NM_001198625	425	runt-related	hsa-miR-	−0.05	0.91
			transcription factor 1;	4458
			translocated to, 1
			(cyclin D-related)
POU2F1	NM_001198783	426	POU class 2	hsa-miR-	−0.02	>0.99
			homeobox 1	4458
A1CF	NM_001198818	427	APOBEC1	hsa-miR-	−0.11	<0.1
			complementation	4500
			factor
ABCC10	NM_001198934	428	ATP-binding cassette,	hsa-miR-	−0.16	0.82	2005,
			sub-family C	4458			2007
			(CFTR/MRP),
			member 10
SMAP2	NM_001198978	429	small ArfGAP2	hsa-miR-	−0.21	0.82	2007,
				4458			2009
AMMECR1L	NM_001199140	430	AMME chromosomal	hsa-let-7d	−0.11	<0.1	2007,
			region gene 1-like				2009
TOR1AIP2	NM_001199260	431	torsin A interacting	hsa-miR-	>−0.03	0.98
			protein 2	4458
UCHL5	NM_001199261	432	ubiquitin carboxyl-	hsa-let-7f	−0.04	<0.1
			terminal hydrolase L5
PHOSPHO2-	NM_001199290	433	PHOSPHO2-	hsa-let-7f	−0.2	0.95
KLHL23			KLHL23 readthrough
CNOT2	NM_001199302	434	CCR4-NOT	hsa-let-7d	−0.14	0.86	2007,
			transcription				2009
			complex, subunit 2
MUTED	NM_001199322	435	muted homolog	hsa-let-7c	−0.1	0.82	2009
			(mouse)
CPD	NM_001199775	436	carboxypeptidase D	hsa-let-7d	−0.18	0.95	2005,
							2007,
							2009
POC1B-	NM_001199781	437	POC1B-GALNT4	hsa-let-7a	−0.1	0.84
GALNT4			readthrough
EGR3	NM_001199880	438	early growth response 3	hsa-miR-	>−0.03	0.67	2005,
				4458			2007,
							2009
DNAL1	NM_001201366	439	dynein, axonemal,	hsa-miR-	−0.03	0.94	2009
			light chain 1	4458
RNF7	NM_001201370	440	ring finger protein 7	hsa-miR-	−0.18	0.94	2005,
				4458			2007,
							2009
TRMT1L	NM_001202423	441	TRM1 tRNA	hsa-miR-	−0.16	0.65
			methyltransferase 1-	4458
			like
HSPE1-	NM_001202485	442	HSPE1-MOBKL3	hsa-let-7a	−0.12	0.93
MOBKL3			readthrough
UBE2G2	NM_001202489	443	ubiquitin-conjugating	hsa-miR-	−0.14	0.94	2009
			enzyme E2G 2	4458
MXD1	NM_001202513	444	MAX dimerization	hsa-miR-	−0.23	0.94	2007,
			protein 1	4458			2009
CUX1	NM_001202543	445	cut-like homeobox 1	hsa-miR-	−0.05	0.77
				4458
SEMA4G	NM_001203244	446	sema domain,	hsa-let-7a	−0.22	0.9	2005,
			immunoglobulin				2007,
			domain (Ig),				2009
			transmembrane
			domain (TM) and
			short cytoplasmic
			domain, (semaphorin)
			4G
EZH2	NM_001203247	447	enhancer of zeste	hsa-miR-	N/A	0.86	2005,
			homolog 2	4458			2007,
			(Drosophila)				2009
PPT2	NM_001204103	448	palmitoyl-protein	hsa-miR-	−0.29	<0.1
			thioesterase 2	4458
MDM4	NM_001204171	449	Mdm4 p53 binding	hsa-let-7a	−0.27	>0.99
			protein homolog
			(mouse)
RGS6	NM_001204416	450	regulator of G-protein	hsa-miR-	−0.22	0.95	2009
			signaling 6	4458
NPEPL1	NM_001204872	451	aminopeptidase-like 1	hsa-let-7f	−0.15	0.94	2005,
							2007,
							2009
PBX1	NM_001204961	452	pre-B-cell leukemia	hsa-let-7g	−0.28	0.94
			homeobox 1
KLF9	NM_001206	453	Kruppel-like factor 9	hsa-miR-	−0.18	0.92	2005,
				4500			2007,
							2009
CD86	NM_001206924	454	CD86 molecule	hsa-miR-	−0.2	0.6	2009
				4500
POU2F2	NM_001207025	455	POU class 2	hsa-miR-	−0.14	0.89	2007,
			homeobox 2	4458			2009
CLASP2	NM_001207044	456	cytoplasmic linker	hsa-let-7g	−0.14	0.94	2005,
			associated protein 2				2007,
							2009
BZW1	NM_001207067	457	basic leucine zipper	hsa-let-7f	−0.48	0.99	2005,
			and W2 domains 1				2007,
							2009
MEIS2	NM_001220482	458	Meis homeobox 2	hsa-miR-	−0.18	0.86	2005,
				4458			2007,
							2009
CCNT2	NM_001241	459	cyclin T2	hsa-miR-	−0.1	0.76
				4458
ATAD2B	NM_001242338	460	ATPase family, AAA	hsa-miR-	>−0.03	0.66	2009
			domain containing 2B	4458
FAM59A	NM_001242409	461	family with sequence	hsa-let-7f	−0.17	0.94
			similarity 59, member A
FBXO32	NM_001242463	462	F-box protein 32	hsa-let-7b	−0.26	0.95
MAP4K4	NM_001242559	463	mitogen-activated	hsa-miR-	−0.2	0.99	2005,
			protein kinase kinase	4458			2007,
			kinase kinase 4				2009
NXT2	NM_001242617	464	nuclear transport	hsa-let-7g	−0.21	0.86	2005,
			factor 2-like export				2007,
			factor 2				2009
GFOD1	NM_001242628	465	glucose-fructose	hsa-miR-	−0.29	<0.1
			oxidoreductase	4458
			domain containing 1
ARHGEF38	NM_001242729	466	Rho guanine	hsa-miR-	−0.53	0.94
			nucleotide exchange	4500
			factor (GEF) 38
ZNF322A	NM_001242797	467	zinc finger protein	hsa-let-7a	−0.47	0.96	2009
			322A
CHD4	NM_001273	468	chromodomain	hsa-miR-	−0.17	0.83	2005,
			helicase DNA binding	4500			2007,
			protein 4				2009
CHUK	NM_001278	469	conserved helix-loop-	hsa-miR-	−0.27	0.74
			helix ubiquitous	4500
			kinase
AP1S1	NM_001283	470	adaptor-related	hsa-miR-	−0.27	0.88	2005,
			protein complex 1,	4458			2007,
			sigma 1 subunit				2009
DUSP4	NM_001394	471	dual specificity	hsa-miR-	−0.17	0.94	2007,
			phosphatase 4	4500			2009
DUSP9	NM_001395	472	dual specificity	hsa-miR-	−0.07	0.93	2005,
			phosphatase 9	4500			2007,
							2009
DYRK1A	NM_001396	473	dual-specificity	hsa-miR-	−0.15	0.84	2005,
			tyrosine-(Y)-	4500			2007,
			phosphorylation				2009
			regulated kinase 1A
GATM	NM_001482	474	glycine	hsa-let-7a	−0.44	0.96	2009
			amidinotransferase
			(L-arginine:glycine
			amidinotransferase)
ACVR2A	NM_001616	475	activin A receptor,	hsa-let-7a	−0.24	0.98	2007,
			type IIA				2009
AKT2	NM_001626	476	v-akt murine	hsa-let-7i	−0.15	0.88	2009
			thymoma viral
			oncogene homolog 2
ARL4D	NM_001661	477	ADP-ribosylation	hsa-miR-	−0.19	0.94	2009
			factor-like 4D	4500
POLR3D	NM_001722	478	polymerase (RNA) III	hsa-miR-	−0.28	0.91	2007,
			(DNA directed)	4500			2009
			polypeptide D, 44 kDa
CCND2	NM_001759	479	cyclin D2	hsa-miR-	−0.14	>0.99	2005,
				4458			2007,
							2009
CCNF	NM_001761	480	cyclin F	hsa-let-7d	−0.4	0.92	2009
CDC25A	NM_001789	481	cell division cycle 25	hsa-miR-	−0.42	0.9	2005,
			homolog A (S. pombe)	4458			2007,
							2009
CCR7	NM_001838	482	chemokine (C-C	hsa-let-7f	−0.36	0.96	2005,
			motif) receptor 7				2007,
							2009
COL4A1	NM_001845	483	collagen, type IV,	hsa-miR-	−0.15	0.92	2005,
			alpha 1	4458			2007,
							2009
COL4A2	NM_001846	484	collagen, type IV,	hsa-miR-98	−0.17	0.94	2005,
			alpha 2				2007,
							2009
COL4A6	NM_001847	485	collagen, type IV,	hsa-miR-	−0.34	<0.1	2009
			alpha 6	4458
COL9A3	NM_001853	486	collagen, type IX,	hsa-let-7a	−0.11	0.86	2009
			alpha 3
COL15A1	NM_001855	487	collagen, type XV,	hsa-miR-	−0.18	0.93	2005,
			alpha 1	4500			2007,
							2009
SLC31A1	NM_001859	488	solute carrier family	hsa-miR-	−0.12	0.94	2009
			31 (copper	4458
			transporters), member 1
SLC31A2	NM_001860	489	solute carrier family	hsa-let-7a	−0.17	0.71	2005,
			31 (copper				2007
			transporters), member 2
MASP1	NM_001879	490	mannan-binding	hsa-let-7a	−0.34	0.94	2007,
			lectin serine peptidase				2009
			1 (C4/C2 activating
			component of Ra-
			reactive factor)
CTPS	NM_001905	491	CTP synthase	hsa-miR-	−0.19	0.79
				4458
DLST	NM_001933	492	dihydrolipoamide S-	hsa-miR-	−0.17	0.95	2005,
			succinyltransferase	4500			2007,
			(E2 component of 2-				2009
			oxo-glutarate
			complex)
DSG3	NM_001944	493	desmoglein 3	hsa-miR-	−0.16	<0.1
				4500
HBEGF	NM_001945	494	heparin-binding EGF-	hsa-miR-	−0.17	0.73
			like growth factor	4500
DUSP7	NM_001947	495	dual specificity	hsa-let-7b	−0.07	0.94
			phosphatase 7
ELK4	NM_001973	496	ELK4, ETS-domain	hsa-miR-	−0.14	0.94
			protein (SRF	4458
			accessory protein 1)
EZH1	NM_001991	497	enhancer of zeste	hsa-let-7a	−0.14	0.88	2009
			homolog 1
			(Drosophila)
GLRX	NM_002064	498	glutaredoxin	hsa-let-7d	−0.23	0.78	2009
			(thioltransferase)
GNS	NM_002076	499	glucosamine (N-	hsa-miR-	>−0.01	0.88	2005,
			acetyl)-6-sulfatase	4458			2007,
							2009
HLF	NM_002126	500	hepatic leukemia	hsa-miR-	−0.06	0.87	2007,
			factor	4500			2009
HMGA1	NM_002131	501	high mobility group	hsa-let-7i	−0.25	0.94	2005,
			AT-hook 1				2007,
							2009
IDH2	NM_002168	502	isocitrate	hsa-let-7d	−0.12	0.83	2005,
			dehydrogenase 2				2007
			(NADP+),
			mitochondrial
IL13	NM_002188	503	interleukin 13	hsa-miR-	−0.32	0.98	2005,
				4500			2007,
							2009
ITGB8	NM_002214	504	integrin, beta 8	hsa-let-7i	−0.1	0.94	2007,
							2009
KCNA6	NM_002235	505	potassium voltage-	hsa-miR-	>−0.03	0.44	2009
			gated channel, shaker-	4458
			related subfamily,
			member 6
KPNA1	NM_002264	506	karyopherin alpha 1	hsa-miR-	−0.15	0.93	2007,
			(importin alpha 5)	4458			2009
KPNA4	NM_002268	507	karyopherin alpha 4	hsa-miR-	−0.08	0.97	2005,
			(importin alpha 3)	4458			2007
LBR	NM_002296	508	lamin B receptor	hsa-miR-	−0.21	0.98	2009
				4458
LY75	NM_002349	509	lymphocyte antigen	hsa-let-7d	−0.17	0.8	2009
			75
MAP3K3	NM_002401	510	mitogen-activated	hsa-miR-	−0.06	0.95	2005,
			protein kinase kinase	4458			2007,
			kinase 3				2009
MSR1	NM_002445	511	macrophage	hsa-let-7a	−0.37	0.98
			scavenger receptor 1
NGF	NM_002506	512	nerve growth factor	hsa-miR-	−0.34	0.93	2009
			(beta polypeptide)	4458
NOVA1	NM_002515	513	neuro-oncological	hsa-let-7d	−0.11	0.92	2005,
			ventral antigen 1				2007,
							2009
NRAS	NM_002524	514	neuroblastoma RAS	hsa-miR-	−0.35	>0.99	2005,
			viral (v-ras) oncogene	4500			2007,
			homolog				2009
P2RX1	NM_002558	515	purinergic receptor	hsa-miR-	−0.12	0.9	2009
			P2X, ligand-gated ion	4458
			channel, 1
PAPPA	NM_002581	516	pregnancy-associated	hsa-miR-98	−0.2	>0.99	2005,
			plasma protein A,				2007,
			pappalysin 1				2009
PBX2	NM_002586	517	pre-B-cell leukemia	hsa-miR-	−0.22	>0.99	2005,
			homeobox 2	4500			2007,
							2009
PDGFB	NM_002608	518	platelet-derived	hsa-miR-	−0.11	0.88	2005,
			growth factor beta	4458			2007,
			polypeptide				2009
PIGA	NM_002641	519	phosphatidylinositol	hsa-miR-	−0.29	0.94	2005,
			glycan anchor	4500			2007,
			biosynthesis, class A				2009
PLAGL2	NM_002657	520	pleiomorphic	hsa-miR-	−0.12	0.91	2005,
			adenoma gene-like 2	4500			2007,
							2009
MAPK6	NM_002748	521	mitogen-activated	hsa-let-7b	−0.38	0.94	2005,
			protein kinase 6				2007,
							2009
MAPK11	NM_002751	522	mitogen-activated	hsa-miR-	−0.11	0.79	2009
			protein kinase 11	4458
MAPK9	NM_002752	523	mitogen-activated	hsa-miR-	−0.05	0.77
			protein kinase 9	4458
PTPRO	NM_002848	524	protein tyrosine	hsa-let-7d	−0.14	0.88	2009
			phosphatase, receptor
			type, O
RALB	NM_002881	525	v-ral simian leukemia	hsa-miR-	−0.17	0.94	2009
			viral oncogene	4458
			homolog B (ras
			related; GTP binding
			protein)
RBMS1	NM_002897	526	RNA binding motif,	hsa-let-7d	−0.25	0.96
			single stranded
			interacting protein 1
RBMS2	NM_002898	527	RNA binding motif,	hsa-let-7a	−0.1	0.98
			single stranded
			interacting protein 2
RCN1	NM_002901	528	reticulocalbin 1, EF-	hsa-let-7d	−0.17	0.85	2009
			hand calcium binding
			domain
RDX	NM_002906	529	radixin	hsa-let-7a	−0.25	0.75	2005,
							2007,
							2009
RGS16	NM_002928	530	regulator of G-protein	hsa-miR-	−0.31	0.98	2005,
			signaling 16	4500			2007,
							2009
S100A8	NM_002964	531	S100 calcium binding	hsa-miR-	−0.18	0.69
			protein A8	4500
CCL3	NM_002983	532	chemokine (C-C	hsa-miR-	−0.23	0.87	2009
			motif) ligand 3	4458
ST3GAL1	NM_003033	533	ST3 beta-galactoside	hsa-miR-	>−0.01	0.89
			alpha-2,3-	4458
			sialyltransferase 1
ST8SIA1	NM_003034	534	ST8 alpha-N-acetyl-	hsa-let-7f	−0.29	0.94	2009
			neuraminide alpha-
			2,8-sialyltransferase 1
SLC6A1	NM_003042	535	solute carrier family 6	hsa-miR-	−0.19	0.95	2005,
			(neurotransmitter	4458			2007,
			transporter, GABA),				2009
			member 1
SMARCC1	NM_003074	536	SWI/SNF related,	hsa-let-7g	−0.13	0.91	2005,
			matrix associated,				2007,
			actin dependent				2009
			regulator of
			chromatin, subfamily
			c, member 1
SNX1	NM_003099	537	sorting nexin 1	hsa-miR-	−0.2	<0.1	2009
				4458
STRN	NM_003162	538	striatin, calmodulin	hsa-miR-	−0.14	0.95
			binding protein	4458
TEAD3	NM_003214	539	TEA domain family	hsa-miR-	−0.08	0.91	2007,
			member 3	4458			2009
THBS1	NM_003246	540	thrombospondin 1	hsa-let-7c	−0.16	0.86	2009
THRSP	NM_003251	541	thyroid hormone	hsa-let-7d	−0.62	0.74	2009
			responsive
VSNL1	NM_003385	542	visinin-like 1	hsa-miR-	−0.11	0.79	2005,
				4458			2007,
							2009
ZNF202	NM_003455	543	zinc finger protein	hsa-let-7d	−0.36	<0.1
			202
BSN	NM_003458	544	bassoon (presynaptic	hsa-let-7a	−0.09	0.94	2009
			cytomatrix protein)
MLL2	NM_003482	545	myeloid/lymphoid or	hsa-miR-	−0.26	0.98	2007,
			mixed-lineage	4458			2009
			leukemia 2
HMGA2	NM_003483	546	high mobility group	hsa-miR-	−1.04	>0.99	2005,
			AT-hook 2	4458			2007,
							2009
SNN	NM_003498	547	stannin	hsa-miR-	−0.15	0.87	2005,
				4458			2007,
							2009
EEA1	NM_003566	548	early endosome	hsa-miR-	−0.21	0.94	2007,
			antigen 1	4458			2009
ZNF282	NM_003575	549	zinc finger protein	hsa-miR-	>−0.02	0.94	2009
			282	4458
DYRK2	NM_003583	550	dual-specificity	hsa-miR-	−0.12	0.87	2009
			tyrosine-(Y)-	4500
			phosphorylation
			regulated kinase 2
SLC4A7	NM_003615	551	solute carrier family	hsa-let-7g	−0.02	0.81	2005,
			4, sodium bicarbonate				2007
			cotransporter,
			member 7
MAP4K3	NM_003618	552	mitogen-activated	hsa-miR-98	−0.46	0.96	2005,
			protein kinase kinase				2007,
			kinase kinase 3				2009
NDST2	NM_003635	553	N-deacetylase/N-	hsa-miR-	−0.32	0.96	2005,
			sulfotransferase	4458			2007,
			(heparan				2009
			glucosaminyl) 2
IKBKAP	NM_003640	554	inhibitor of kappa	hsa-miR-	−0.27	0.93	2005,
			light polypeptide gene	4458			2007,
			enhancer in B-cells,				2009
			kinase complex-
			associated protein
CHRD	NM_003741	555	chordin	hsa-miR-	−0.15	0.87	2005,
				4458			2007
NCOA1	NM_003743	556	nuclear receptor	hsa-let-7d	−0.07	0.7	2005,
			coactivator 1				2007
IRS2	NM_003749	557	insulin receptor	hsa-miR-	−0.28	0.98	2005,
			substrate 2	4458			2007,
							2009
GALNT4	NM_003774	558	UDP-N-acetyl-alpha-	hsa-let-7a	−0.1	0.84	2009
			D-
			galactosamine:polypeptide
			N-
			acetylgalactosaminyltransferase 4
			(GalNAc-T4)
RNMT	NM_003799	559	RNA (guanine-7-)	hsa-miR-	−0.31	<0.1
			methyltransferase	4458
TNFSF9	NM_003811	560	tumor necrosis factor	hsa-let-7d	−0.35	0.98	2009
			(ligand) superfamily,
			member 9
SNAP23	NM_003825	561	synaptosomal-	hsa-miR-	−0.26	0.93	2005,
			associated protein,	4500			2007,
			23 kDa				2009
RIOK3	NM_003831	562	RIO kinase 3 (yeast)	hsa-miR-	−0.26	0.94	2005,
				4500			2007,
							2009
SUCLG2	NM_003848	563	succinate-CoA ligase,	hsa-let-7a	−0.14	0.89	2009
			GDP-forming, beta
			subunit
EIF2S2	NM_003908	564	eukaryotic translation	hsa-miR-	−0.22	0.92	2009
			initiation factor 2,	4458
			subunit 2 beta, 38 kDa
MBD2	NM_003927	565	methyl-CpG binding	hsa-let-7f	−0.29	>0.99
			domain protein 2
WASL	NM_003941	566	Wiskott-Aldrich	hsa-let-7d	−0.17	0.87	2009
			syndrome-like
RNF8	NM_003958	567	ring finger protein 8	hsa-miR-	−0.1	0.68
				4500
OSMR	NM_003999	568	oncostatin M receptor	hsa-miR-	−0.27	0.94	2005,
				4458			2007,
							2009
E2F2	NM_004091	569	E2F transcription	hsa-let-7d	−0.28	0.94	2009
			factor 2
FGF11	NM_004112	570	fibroblast growth	hsa-miR-98	−0.26	0.93	2005,
			factor 11				2007,
							2009
TARBP2	NM_004178	571	TAR (HIV-1) RNA	hsa-miR-	−0.2	0.93	2009
			binding protein 2	4458
SEMA3F	NM_004186	572	sema domain,	hsa-miR-	−0.16	0.72	2005,
			immunoglobulin	4458			2007
			domain (Ig), short
			basic domain,
			secreted,
			(semaphorin) 3F
SYT7	NM_004200	573	synaptotagmin VII	hsa-miR-98	−0.25	0.98	2007,
							2009
AURKB	NM_004217	574	aurora kinase B	hsa-miR-	−0.26	0.78
				4458
CYTH3	NM_004227	575	cytohesin 3	hsa-miR-	−0.02	0.9	2005,
				4458			2007,
							2009
SEMA4F	NM_004263	576	sema domain,	hsa-miR-	−0.4	0.97	2009
			immunoglobulin	4500
			domain (Ig),
			transmembrane
			domain (TM) and
			short cytoplasmic
			domain, (semaphorin)
			4F
CHST3	NM_004273	577	carbohydrate	hsa-miR-	−0.11	0.99	2009
			(chondroitin 6)	4458
			sulfotransferase 3
AKAP6	NM_004274	578	A kinase (PRKA)	hsa-let-7c	−0.28	0.95	2005,
			anchor protein 6				2007,
							2009
SLC25A27	NM_004277	579	solute carrier family	hsa-miR-	−0.29	0.95	2005,
			25, member 27	4458			2007,
							2009
ACVR1B	NM_004302	580	activin A receptor,	hsa-let-7g	−0.12	0.95	2005,
			type IB				2007,
							2009
CASP3	NM_004346	581	caspase 3, apoptosis-	hsa-let-7b	−0.4	0.99	2005,
			related cysteine				2007,
			peptidase				2009
CDC34	NM_004359	582	cell division cycle 34	hsa-miR-	−0.47	>0.99	2005,
			homolog (S. cerevisiae)	4500			2007,
							2009
DUSP1	NM_004417	583	dual specificity	hsa-miR-	−0.18	0.87	2005,
			phosphatase 1	4500			2007,
							2009
DVL3	NM_004423	584	dishevelled, dsh	hsa-let-7f	−0.41	0.89	2009
			homolog 3
			(Drosophila)
EPHA4	NM_004438	585	EPH receptor A4	hsa-miR-	−0.18	0.87	2005,
				4500			2007,
							2009
FGF5	NM_004464	586	fibroblast growth	hsa-miR-	−0.11	0.93	2009
			factor 5	4458
GALNT2	NM_004481	587	UDP-N-acetyl-alpha-	hsa-miR-	−0.18	0.94	2005,
			D-	4500			2007,
			galactosamine:polypeptide				2009
			N-
			acetylgalactosaminyltransferase 2
			(GalNAc-T2)
USP6	NM_004505	588	ubiquitin specific	hsa-let-7a	−0.18	0.92	2005,
			peptidase 6 (Tre-2				2007,
			oncogene)				2009
NAP1L1	NM_004537	589	nucleosome assembly	hsa-let-7d	−0.41	>0.99	2005,
			protein 1-like 1				2007,
							2009
NRTN	NM_004558	590	neurturin	hsa-miR-	N/A	0.85	2007,
				4458			2009
RPS6KA3	NM_004586	591	ribosomal protein S6	hsa-miR-98	−0.1	0.94	2005,
			kinase, 90 kDa,				2007,
			polypeptide 3				2009
COIL	NM_004645	592	coilin	hsa-miR-	−0.5	0.96	2005,
				4500			2007,
							2009
KCNQ4	NM_004700	593	potassium voltage-	hsa-let-7d	−0.18	0.94
			gated channel, KQT-
			like subfamily,
			member 4
NUMBL	NM_004756	594	numb homolog	hsa-miR-	−0.04	0.94	2005,
			(Drosophila)-like	4500			2007,
							2009
NDST3	NM_004784	595	N-deacetylase/N-	hsa-let-7d	−0.18	0.83
			sulfotransferase
			(heparan
			glucosaminyl) 3
POLR2D	NM_004805	596	polymerase (RNA) II	hsa-let-7b	−0.42	<0.1	2009
			(DNA directed)
			polypeptide D
HAND1	NM_004821	597	heart and neural crest	hsa-let-7a	−0.44	0.98	2005,
			derivatives expressed 1				2007,
							2009
NTN1	NM_004822	598	netrin 1	hsa-miR-	−0.26	0.84	2009
				4500
ONECUT2	NM_004852	599	one cut homeobox 2	hsa-miR-	−0.13	>0.99	2007,
				4500			2009
AKAP5	NM_004857	600	A kinase (PRKA)	hsa-miR-	>−0.03	0.72	2009
			anchor protein 5	4458
IGDCC3	NM_004884	601	immunoglobulin	hsa-miR-	−0.72	>0.99	2003,
			superfamily, DCC	4500			2007,
			subclass, member 3				2009
SEC24C	NM_004922	602	SEC24 family,	hsa-miR-98	−0.1	0.82	2005,
			member C (S. cerevisiae)				2007,
							2009
DAPK1	NM_004938	603	death-associated	hsa-let-7g	−0.18	0.93	2009
			protein kinase 1
DOCK3	NM_004947	604	dedicator of	hsa-miR-	−0.07	0.9	2005,
			cytokinesis 3	4458			2007,
							2009
KCNC1	NM_004976	605	potassium voltage-	hsa-miR-	−0.1	0.91	2009
			gated channel, Shaw-	4458
			related subfamily,
			member 1
NME4	NM_005009	606	non-metastatic cells 4,	hsa-let-7d	−0.19	0.94	2003,
			protein expressed in				2005,
							2007,
							2009
QARS	NM_005051	607	glutaminyl-tRNA	hsa-let-7i	−0.37	0.2	2005,
			synthetase				2007,
							2009
SCD	NM_005063	608	stearoyl-CoA	hsa-miR-98	−0.33	0.95	2007,
			desaturase (delta-9-				2009
			desaturase)
SIM2	NM_005069	609	single-minded	hsa-let-7d	−0.08	0.9	2009
			homolog 2
			(Drosophila)
ADRBK2	NM_005160	610	adrenergic, beta,	hsa-miR-98	−0.15	0.89	2009
			receptor kinase 2
CBFA2T3	NM_005187	611	core-binding factor,	hsa-miR-	−0.06	0.94	2005,
			runt domain, alpha	4458			2007,
			subunit 2;				2009
			translocated to, 3
CBL	NM_005188	612	Cas-Br-M (murine)	hsa-miR-	−0.07	0.96	2005,
			ecotropic retroviral	4500			2007,
			transforming				2009
			sequence
CBX2	NM_005189	613	chromobox homolog 2	hsa-miR-	−0.15	0.94	2007,
				4458			2009
CEBPD	NM_005195	614	CCAAT/enhancer	hsa-let-7d	−0.09	0.86	2009
			binding protein
			(C/EBP), delta
ARID3A	NM_005224	615	AT rich interactive	hsa-miR-	−0.11	0.9	2007,
			domain 3A	4458			2009
			(BRIGHT-like)
EPHA3	NM_005233	616	EPH receptor A3	hsa-miR-	−0.14	0.93	2005,
				4500			2007
ERCC4	NM_005236	617	excision repair cross-	hsa-miR-	−0.1	0.98	2009
			complementing	4500
			rodent repair
			deficiency,
			complementation
			group
4
GNG5	NM_005274	618	guanine nucleotide	hsa-miR-	−0.28	0.81	2005,
			binding protein (G	4458			2007,
			protein), gamma 5				2009
HAS2	NM_005328	619	hyaluronan synthase 2	hsa-let-7d	−0.12	0.87	2005,
							2007,
							2009
HDLBP	NM_005336	620	high density	hsa-miR-	−0.21	0.98	2007,
			lipoprotein binding	4458			2009
			protein
MYCN	NM_005378	621	v-myc	hsa-let-7i	−0.34	0.95	2005,
			myelocytomatosis				2007,
			viral related				2009
			oncogene,
			neuroblastoma
			derived (avian)
NUP98	NM_005387	622	nucleoporin 98 kDa	hsa-let-7d	−0.23	0.85
PRKAB2	NM_005399	623	protein kinase, AMP-	hsa-miR-	>−0.02	0.89	2009
			activated, beta 2 non-	4458
			catalytic subunit
SLC20A1	NM_005415	624	solute carrier family	hsa-miR-	−0.31	0.87	2005,
			20 (phosphate	4458			2007
			transporter), member 1
WNT1	NM_005430	625	wingless-type MMTV	hsa-let-7d	−0.07	0.88	2005,
			integration site				2007,
			family, member 1				2009
GABBR2	NM_005458	626	gamma-aminobutyric	hsa-miR-	−0.07	0.89	2009
			acid (GABA) B	4458
			receptor, 2
MED6	NM_005466	627	mediator complex	hsa-let-7d	−0.14	0.86	2005,
			subunit 6				2007,
							2009
SH2B3	NM_005475	628	SH2B adaptor protein 3	hsa-miR-	−0.13	0.94	2007,
				4500			2009
INPP5A	NM_005539	629	inositol	hsa-let-7d	−0.1	0.92	2005,
			polyphosphate-5-				2007,
			phosphatase, 40 kDa				2009
ISLR	NM_005545	630	immunoglobulin	hsa-let-7b	−0.23	0.86
			superfamily
			containing leucine-
			rich repeat
LIMK2	NM_005569	631	LIM domain kinase 2	hsa-miR-98	−0.14	0.76
MDFI	NM_005586	632	MyoD family	hsa-miR-	−0.11	0.94	2007,
			inhibitor	4500			2009
ZNF354A	NM_005649	633	zinc finger protein	hsa-miR-	−0.27	0.93	2005,
			354A	4500			2007,
							2009
SOX13	NM_005686	634	SRY (sex determining	hsa-let-7g	−0.16	0.9	2005,
			region Y)-box 13				2007,
							2009
ABCC5	NM_005688	635	ATP-binding cassette,	hsa-miR-	−0.34	0.67	2005,
			sub-family C	4500			2007,
			(CFTR/MRP),				2009
			member 5
DPP3	NM_005700	636	dipeptidyl-peptidase 3	hsa-miR-	−0.35	0.91	2005,
				4458			2007,
							2009
GIPC1	NM_005716	637	GIPC PDZ domain	hsa-miR-	−0.26	0.9	2007,
			containing family,	4458			2009
			member 1
TSPAN2	NM_005725	638	tetraspanin 2	hsa-let-7g	−0.09	0.94	2009
PLXNC1	NM_005761	639	plexin C1	hsa-miR-	−0.33	0.33
				4458
AASS	NM_005763	640	aminoadipate-	hsa-miR-	−0.16	<0.1
			semialdehyde	4458
			synthase
FARP1	NM_005766	641	FERM, RhoGEF	hsa-miR-	−0.16	0.93	2005,
			(ARHGEF) and	4500			2007,
			pleckstrin domain				2009
			protein 1
			(chondrocyte-derived)
NME6	NM_005793	642	non-metastatic cells 6,	hsa-miR-	−0.27	0.94	2005,
			protein expressed in	4458			2007,
			(nucleoside-				2009
			diphosphate kinase)
SPEG	NM_005876	643	SPEG complex locus	hsa-let-7d	−0.11	0.9	2009
DNAJA2	NM_005880	644	DnaJ (Hsp40)	hsa-miR-	−0.28	0.79
			homolog, subfamily	4458
			A, member 2
APC2	NM_005883	645	adenomatosis	hsa-miR-	−0.11	0.83	2009
			polyposis coli 2	4458
MEF2D	NM_005920	646	myocyte enhancer	hsa-miR-	>−0.04	0.94	2005,
			factor 2D	4458			2007,
							2009
MAP3K1	NM_005921	647	mitogen-activated	hsa-miR-	−0.39	0.86	2009
			protein kinase kinase	4458
			kinase 1
MMP11	NM_005940	648	matrix	hsa-let-7d	−0.12	0.92	2007,
			metallopeptidase 11				2009
			(stromelysin 3)
ALKBH1	NM_006020	649	alkB, alkylation repair	hsa-miR-	−0.16	0.89	2009
			homolog 1 (E. coli)	4500
APBB3	NM_006051	650	amyloid beta (A4)	hsa-let-7a	−0.41	0.98	2005,
			precursor protein-				2007,
			binding, family B,				2009
			member 3
DAGLA	NM_006133	651	diacylglycerol lipase,	hsa-miR-	−0.27	0.97	2007,
			alpha	4458			2009
PRKAA2	NM_006252	652	protein kinase, AMP-	hsa-let-7f	−0.17	0.93	2009
			activated, alpha 2
			catalytic subunit
RANBP2	NM_006267	653	RAN binding protein 2	hsa-miR-	−0.38	0.98	2005,
				4458			2007,
							2009
DPF2	NM_006268	654	D4, zinc and double	hsa-let-7g	−0.21	0.94	2005,
			PHD fingers family 2				2007,
							2009
CCL7	NM_006273	655	chemokine (C-C	hsa-miR-	−0.39	0.9	2009
			motif) ligand 7	4458
TNFAIP3	NM_006290	656	tumor necrosis factor,	hsa-miR-	−0.08	0.92	2009
			alpha-induced protein 3	4458
SMC1A	NM_006306	657	structural	hsa-let-7a	−0.46	>0.99	2009
			maintenance of
			chromosomes 1A
PCGF3	NM_006315	658	polycomb group ring	hsa-let-7a	−0.11	0.98	2005,
			finger 3				2007,
							2009
CRTAP	NM_006371	659	cartilage associated	hsa-miR-	−0.11	0.94	2005,
			protein	4500			2007,
							2009
APPBP2	NM_006380	660	amyloid beta	hsa-miR-	−0.02	0.92	2005,
			precursor protein	4458			2007
			(cytoplasmic tail)
			binding protein 2
OLFM4	NM_006418	661	olfactomedin 4	hsa-miR-	−0.37	<0.1
				4500
ARID3B	NM_006465	662	AT rich interactive	hsa-let-7i	−0.72	>0.99	2005,
			domain 3B				2007,
			(BRIGHT-like)				2009
IGF2BP3	NM_006547	663	insulin-like growth	hsa-let-7a	−0.33	0.96	2007,
			factor 2 mRNA				2009
			binding protein 3
CLDN16	NM_006580	664	claudin 16	hsa-miR-98	−0.3	<0.1
MAP3K2	NM_006609	665	mitogen-activated	hsa-let-7a	−0.1	0.8
			protein kinase kinase
			kinase
2
ARPP19	NM_006628	666	cAMP-regulated	hsa-miR-98	−0.09	0.98	2005,
			phosphoprotein,				2007,
			19 kDa				2009
PGRMC1	NM_006667	667	progesterone receptor	hsa-miR-	−0.46	0.98	2005,
			membrane component 1	4458			2007,
							2009
CYP46A1	NM_006668	668	cytochrome P450,	hsa-let-7a	−0.17	0.89	2007,
			family 46, subfamily				2009
			A, polypeptide 1
SUB1	NM_006713	669	SUB1 homolog (S. cerevisiae)	hsa-miR-	−0.36	0.84
				4500
BTG2	NM_006763	670	BTG family, member 2	hsa-miR-	−0.12	0.89	2005,
				4458			2007,
							2009
PKIA	NM_006823	671	protein kinase	hsa-miR-	−0.18	0.94
			(cAMP-dependent,	4458
			catalytic) inhibitor
			alpha
B3GNT1	NM_006876	672	UDP-	hsa-miR-	−0.34	0.93	2007,
			GlcNAc:betaGal beta-	4500			2009
			1,3-N-
			acetylglucosaminyltransferase 1
CALM1	NM_006888	673	calmodulin 1	hsa-miR-	−0.1	0.93	2005,
			(phosphorylase	4500			2007,
			kinase, delta)				2009
PRRX1	NM_006902	674	paired related	hsa-miR-98	−0.05	0.95	2007,
			homeobox 1				2009
RNF5	NM_006913	675	ring finger protein 5	hsa-miR-	−0.26	0.64	2005,
				4458			2007,
							2009
ZNF24	NM_006965	676	zinc finger protein 24	hsa-miR-	−0.12	0.94
				4500
ADAMTS1	NM_006988	677	ADAM	hsa-miR-98	−0.12	0.84	2009
			metallopeptidase with
			thrombospondin type
			1 motif, 1
ZNF197	NM_006991	678	zinc finger protein	hsa-let-7a	−0.38	0.12
			197
SLC35D2	NM_007001	679	solute carrier family	hsa-let-7d	−0.48	0.98	2005,
			35, member D2				2007,
							2009
CNTRL	NM_007018	680	centriolin	hsa-miR-	−0.42	0.98	2009
				4458
ADAMTS8	NM_007037	681	ADAM	hsa-miR-	−0.4	0.98	2007,
			metallopeptidase with	4500			2009
			thrombospondin type
			1 motif, 8
ADAMTS5	NM_007038	682	ADAM	hsa-miR-	−0.18	0.95	2005,
			metallopeptidase with	4458			2007,
			thrombospondin type				2009
			1 motif, 5
UTRN	NM_007124	683	utrophin	hsa-miR-	−0.35	0.9	2007,
				4458			2009
ZNF81	NM_007137	684	zinc finger protein 81	hsa-miR-	−0.07	0.81
				4500
TUSC2	NM_007275	685	tumor suppressor	hsa-miR-	−0.12	0.93	2005,
			candidate 2	4458			2007,
							2009
AP4E1	NM_007347	686	adaptor-related	hsa-miR-	>−0.02	0.57
			protein complex 4,	4458
			epsilon 1 subunit
NID2	NM_007361	687	nidogen 2	hsa-miR-	−0.16	0.92	2005,
			(osteonidogen)	4500			2007,
							2009
BRD3	NM_007371	688	bromodomain	hsa-miR-	−0.11	0.94	2005,
			containing 3	4500			2007,
							2009
ICOS	NM_012092	689	inducible T-cell co-	hsa-let-7b	−0.24	0.92	2009
			stimulator
ANGPTL2	NM_012098	690	angiopoietin-like 2	hsa-miR-	−0.13	0.93	2005,
				4458			2007,
							2009
BACE2	NM_012105	691	beta-site APP-	hsa-miR-	−0.27	0.93	2009
			cleaving enzyme 2	4500
FZD4	NM_012193	692	frizzled family	hsa-miR-	−0.33	0.94	2007,
			receptor 4	4500			2009
EIF2C1	NM_012199	693	eukaryotic translation	hsa-miR-	−0.16	0.93	2005,
			initiation factor 2C, 1	4500			2007,
							2009
B3GAT3	NM_012200	694	beta-1,3-	hsa-miR-	−0.18	0.88	2009
			glucuronyltransferase 3	4458
			(glucuronosyltransferase
			I)
MGAT4A	NM_012214	695	mannosyl (alpha-1,3-)-	hsa-miR-	−0.29	0.95	2005,
			glycoprotein beta-	4500			2007,
			1,4-N-				2009
			acetylglucosaminyltransferase,
			isozyme A
HS2ST1	NM_012262	696	heparan sulfate 2-O-	hsa-miR-98	−0.06	0.89	2009
			sulfotransferase 1
ESPL1	NM_012291	697	extra spindle pole	hsa-miR-98	−0.28	0.57
			bodies homolog 1 (S. cerevisiae)
PXDN	NM_012293	698	peroxidasin homolog	hsa-miR-	−0.12	>0.99	2007,
			(Drosophila)	4500			2009
GAB2	NM_012296	699	GRB2-associated	hsa-miR-	−0.15	0.6	2009
			binding protein 2	4458
PLA2G15	NM_012320	700	phospholipase A2,	hsa-miR-	−0.09	0.92	2005,
			group XV	4458			2007,
							2009
DNAJB9	NM_012328	701	DnaJ (Hsp40)	hsa-miR-	−0.1	0.91	2005,
			homolog, subfamily	4500			2007,
			B, member 9				2009
MYCBP	NM_012333	702	c-myc binding protein	hsa-miR-	−0.08	0.98	2009
				4458
MYO1F	NM_012335	703	myosin IF	hsa-miR-	−0.22	0.93	2007,
				4500			2009
NNT	NM_012343	704	nicotinamide	hsa-miR-	−0.26	0.87	2009
			nucleotide	4458
			transhydrogenase
PLDN	NM_012388	705	pallidin homolog	hsa-miR-	−0.07	0.94	2005,
			(mouse)	4500			2007,
							2009
CDK14	NM_012395	706	cyclin-dependent	hsa-miR-	−0.16	0.67
			kinase 14	4500
ICMT	NM_012405	707	isoprenylcysteine	hsa-miR-	>−0.03	0.98	2009
			carboxyl	4458
			methyltransferase
RAB3GAP2	NM_012414	708	RAB3 GTPase	hsa-let-7d	−0.18	0.94
			activating protein
			subunit 2 (non-
			catalytic)
PPARGC1A	NM_013261	709	peroxisome	hsa-miR-	−0.11	0.83	2005,
			proliferator-activated	4500			2007,
			receptor gamma,				2009
			coactivator 1 alpha
EEF2K	NM_013302	710	eukaryotic elongation	hsa-let-7d	−0.17	0.95	2007,
			factor-2 kinase				2009
SLC30A4	NM_013309	711	solute carrier family	hsa-let-7a	−0.14	0.94	2005,
			30 (zinc transporter),				2007,
			member 4				2009
HCFC2	NM_013320	712	host cell factor C2	hsa-miR-	−0.04	0.91
				4500
ATG4B	NM_013325	713	ATG4 autophagy	hsa-let-7a	−0.19	0.79	2009
			related 4 homolog B
			(S. cerevisiae)
GPR132	NM_013345	714	G protein-coupled	hsa-let-7f	−0.21	<0.1
			receptor 132
TRHDE	NM_013381	715	thyrotropin-releasing	hsa-let-7d	−0.19	0.99	2005,
			hormone degrading				2007,
			enzyme				2009
SLC25A24	NM_013386	716	solute carrier family	hsa-miR-	−0.24	0.94	2005,
			25 (mitochondrial	4458			2007,
			carrier; phosphate				2009
			carrier), member 24
WDR37	NM_014023	717	WD repeat domain 37	hsa-let-7a	−0.21	0.99	2005,
							2007,
							2009
PSORS1C2	NM_014069	718	psoriasis	hsa-let-7d	−0.15	0.93
			susceptibility 1
			candidate 2
SCN11A	NM_014139	719	sodium channel,	hsa-miR-	−0.36	0.91	2007,
			voltage-gated, type	4458			2009
			XI, alpha subunit
HOXC11	NM_014212	720	homeobox C11	hsa-miR-	−0.08	0.94	2005,
				4458			2007,
							2009
LIMD1	NM_014240	721	LIM domains	hsa-let-7b	−0.05	0.92	2005,
			containing 1				2007,
							2009
HABP4	NM_014282	722	hyaluronan binding	hsa-miR-	−0.18	0.94	2009
			protein 4	4458
TGDS	NM_014305	723	TDP-glucose 4,6-	hsa-miR-	−0.24	0.95	2009
			dehydratase	4500
SMUG1	NM_014311	724	single-strand-	hsa-let-7d	−0.35	0.92	2009
			selective
			monofunctional
			uracil-DNA
			glycosylase
1
CACNG4	NM_014405	725	calcium channel,	hsa-miR-	−0.17	0.84	2005,
			voltage-dependent,	4458			2007,
			gamma subunit 4				2009
KIAA1274	NM_014431	726	KIAA1274	hsa-miR-98	−0.38	0.98	2009
ZKSCAN5	NM_014569	727	zinc finger with	hsa-miR-	−0.1	0.77
			KRAB and SCAN	4458
			domains 5
ERO1L	NM_014584	728	ERO1-like (S. cerevisiae)	hsa-miR-	−0.18	0.88	2009
				4500
SOCS7	NM_014598	729	suppressor of	hsa-miR-	>−0.02	0.92
			cytokine signaling 7	4458
UBXN4	NM_014607	730	UBX domain protein 4	hsa-let-7a	−0.15	0.89	2005,
							2007,
							2009
RALGPS1	NM_014636	731	Ral GEF with PH	hsa-miR-	−0.15	0.94	2005,
			domain and SH3	4500			2007,
			binding motif 1				2009
TTLL4	NM_014640	732	tubulin tyrosine	hsa-let-7d	−0.56	>0.99	2003,
			ligase-like family,				2005,
			member 4				2007,
							2009
ZNF516	NM_014643	733	zinc finger protein	hsa-miR-	−0.04	0.95	2009
			516	4500
GREB1	NM_014668	734	growth regulation by	hsa-miR-	−0.17	0.88	2009
			estrogen in breast	4500
			cancer 1
ULK2	NM_014683	735	unc-51-like kinase 2	hsa-miR-98	−0.14	0.95	2005,
			(C. elegans)				2007,
							2009
SEC14L5	NM_014692	736	SEC14-like 5 (S. cerevisiae)	hsa-let-7d	−0.14	0.98	2009
TBKBP1	NM_014726	737	TBK1 binding protein 1	hsa-miR-	−0.36	0.97	2007,
				4500			2009
RIMS3	NM_014747	738	regulating synaptic	hsa-miR-	>−0.01	0.92	2009
			membrane exocytosis 3	4458
TSC22D2	NM_014779	739	TSC22 domain	hsa-miR-	−0.16	0.87	2005,
			family, member 2	4458			2007,
							2009
LRIG2	NM_014813	740	leucine-rich repeats	hsa-let-7d	−0.48	0.95	2005,
			and immunoglobulin-				2007,
			like domains 2				2009
ZBTB39	NM_014830	741	zinc finger and BTB	hsa-miR-	−0.04	0.98	2007,
			domain containing 39	4458			2009
TRANK1	NM_014831	742	tetratricopeptide	hsa-let-7b	−0.3	0.85	2009
			repeat and ankyrin
			repeat containing 1
TECPR2	NM_014844	743	tectonin beta-	hsa-let-7d	−0.13	0.94	2005,
			propeller repeat				2007,
			containing 2				2009
ZBTB5	NM_014872	744	zinc finger and BTB	hsa-let-7f	−0.23	0.92	2005,
			domain containing 5				2007,
							2009
LPGAT1	NM_014873	745	lysophosphatidylglycerol	hsa-let-7g	−0.34	>0.99	2005,
			acyltransferase 1				2007,
							2009
HELZ	NM_014877	746	helicase with zinc	hsa-let-7d	−0.28	0.9
			finger
RNF44	NM_014901	747	ring finger protein 44	hsa-let-7i	−0.14	0.94	2005,
							2007,
							2009
AAK1	NM_014911	748	AP2 associated kinase 1	hsa-miR-	>−0.03	0.92	2007,
				4458			2009
DZIP1	NM_014934	749	DAZ interacting	hsa-miR-	−0.11	0.94	2005,
			protein 1	4500			2007,
							2009
MLXIP	NM_014938	750	MLX interacting	hsa-miR-	−0.11	0.8
			protein	4458
BAHD1	NM_014952	751	bromo adjacent	hsa-miR-	−0.05	0.89	2005,
			homology domain	4458			2007,
			containing 1				2009
CEP164	NM_014956	752	centrosomal protein	hsa-miR-	−0.08	0.93	2005,
			164 kDa	4458			2007,
							2009
RUFY3	NM_014961	753	RUN and FYVE	hsa-let-7f	−0.13	0.93	2007,
			domain containing 3				2009
BTBD3	NM_014962	754	BTB (POZ) domain	hsa-let-7c	−0.19	0.92	2005,
			containing 3				2007,
							2009
MON2	NM_015026	755	MON2 homolog (S. cerevisiae)	hsa-miR-	−0.09	0.94	2007,
				4458			2009
NMNAT2	NM_015039	756	nicotinamide	hsa-miR-	>−0.01	0.72
			nucleotide	4458
			adenylyltransferase 2
RRP1B	NM_015056	757	ribosomal RNA	hsa-miR-	−0.09	0.93	2007,
			processing 1 homolog	4458			2009
			B (S. cerevisiae)
SLC8A2	NM_015063	758	solute carrier family 8	hsa-miR-	−0.07	0.91	2005,
			(sodium/calcium	4458			2007,
			exchanger), member 2				2009
DDN	NM_015086	759	dendrin	hsa-miR-	−0.18	0.92	2009
				4500
TAB2	NM_015093	760	TGF-beta activated	hsa-miR-	−0.16	0.83	2005,
			kinase 1/MAP3K7	4500			2007,
			binding protein 2				2009
HIC2	NM_015094	761	hypermethylated in	hsa-miR-	−0.45	>0.99	2003,
			cancer 2	4458			2005,
							2007,
							2009
PLXND1	NM_015103	762	plexin D1	hsa-miR-	−0.35	0.98	2007,
				4500			2009
ZC3H3	NM_015117	763	zinc finger CCCH-	hsa-let-7d	−0.3	0.98	2007,
			type containing 3				2009
FRMD4B	NM_015123	764	FERM domain	hsa-miR-	−0.31	0.86	2009
			containing 4B	4500
DTX4	NM_015177	765	deltex homolog 4	hsa-miR-	−0.06	0.98	2009
			(Drosophila)	4500
OTUD3	NM_015207	766	OTU domain	hsa-miR-	>−0.01	0.82	2009
			containing 3	4458
KHNYN	NM_015299	767	KH and NYN domain	hsa-let-7f	−0.44	0.93
			containing
USP24	NM_015306	768	ubiquitin specific	hsa-miR-	−0.26	0.93	2009
			peptidase 24	4458
FAM189A1	NM_015307	769	family with sequence	hsa-miR-	−0.19	0.94	2009
			similarity 189,	4458
			member A1
LEPROTL1	NM_015344	770	leptin receptor	hsa-let-7d	−0.09	0.82	2005,
			overlapping				2007
			transcript-like 1
ZFYVE26	NM_015346	771	zinc finger, FYVE	hsa-miR-	−0.38	0.98	2005,
			domain containing 26	4458			2007,
							2009
PARM1	NM_015393	772	prostate androgen-	hsa-miR-	−0.24	0.97	2009
			regulated mucin-like	4458
			protein 1
ARMC8	NM_015396	773	armadillo repeat	hsa-miR-	−0.1	0.87
			containing 8	4500
AHCTF1	NM_015446	774	AT hook containing	hsa-miR-	−0.38	0.4	2007,
			transcription factor 1	4458			2009
MYRIP	NM_015460	775	myosin VIIA and Rab	hsa-miR-	−0.11	0.85	2005,
			interacting protein	4458			2007,
							2009
SLC22A23	NM_015482	776	solute carrier family	hsa-miR-	−0.27	0.98
			22, member 23	4458
PNKD	NM_015488	777	paroxysmal	hsa-miR-	−0.14	0.76	2007,
			nonkinesigenic	4458			2009
			dyskinesia
SEC31B	NM_015490	778	SEC31 homolog B (S. cerevisiae)	hsa-miR-	−0.15	0.82	2009
				4458
C15orf39	NM_015492	779	chromosome 15 open	hsa-let-7a	−0.31	0.93	2009
			reading frame 39
LRIG1	NM_015541	780	leucine-rich repeats	hsa-miR-	N/A	0.95	2005,
			and immunoglobulin-	4458			2007,
			like domains 1				2009
OSBPL3	NM_015550	781	oxysterol binding	hsa-miR-	−0.2	0.95	2005,
			protein-like 3	4458			2007,
							2009
LTN1	NM_015565	782	listerin E3 ubiquitin	hsa-let-7d	−0.22	0.93	2005,
			protein ligase 1				2007,
							2009
PLA2G3	NM_015715	783	phospholipase A2,	hsa-let-7a	−0.35	0.91	2009
			group III
DCAF8	NM_015726	784	DDB1 and CUL4	hsa-miR-	−0.1	0.66
			associated factor 8	4500
WARS2	NM_015836	785	tryptophanyl tRNA	hsa-miR-	−0.3	<0.1	2009
			synthetase 2,	4458
			mitochondrial
MBTPS2	NM_015884	786	membrane-bound	hsa-miR-	−0.06	0.86
			transcription factor	4458
			peptidase, site 2
HOOK1	NM_015888	787	hook homolog 1	hsa-miR-	−0.12	0.94	2007,
			(Drosophila)	4458			2009
TAF9B	NM_015975	788	TAF9B RNA	hsa-let-7a	−0.3	0.98	2009
			polymerase II, TATA
			box binding protein
			(TBP)-associated
			factor, 31 kDa
GOLT1B	NM_016072	789	golgi transport 1B	hsa-miR-	−0.18	0.98	2005,
				4500			2007,
							2009
CERCAM	NM_016174	790	cerebral endothelial	hsa-let-7d	−0.15	0.93	2007,
			cell adhesion				2009
			molecule
VGLL3	NM_016206	791	vestigial like 3	hsa-miR-	−0.11	0.94	2009
			(Drosophila)	4458
NLK	NM_016231	792	nemo-like kinase	hsa-miR-	−0.14	0.87	2005,
				4500			2007,
							2009
SCARA3	NM_016240	793	scavenger receptor	hsa-miR-	−0.03	0.77
			class A, member 3	4500
IMPG2	NM_016247	794	interphotoreceptor	hsa-let-7i	−0.31	0.95
			matrix proteoglycan 2
TOB2	NM_016272	795	transducer of ERBB2, 2	hsa-miR-	>−0.02	0.94	2005,
				4458			2007,
							2009
PLEKHO1	NM_016274	796	pleckstrin homology	hsa-miR-	−0.2	0.79	2007,
			domain containing,	4458			2009
			family O member 1
ANKFY1	NM_016376	797	ankyrin repeat and	hsa-miR-	>−0.03	0.98	2005,
			FYVE domain	4458			2007,
			containing 1				2009
LUC7L3	NM_016424	798	LUC7-like 3 (S. cerevisiae)	hsa-miR-	−0.05	0.85	2005,
				4500			2007,
							2009
RAB8B	NM_016530	799	RAB8B, member	hsa-let-7f	−0.07	0.92	2007
			RAS oncogene family
GCNT4	NM_016591	800	glucosaminyl (N-	hsa-miR-	−0.34	0.91	2007,
			acetyl) transferase 4,	4500			2009
			core 2
UFM1	NM_016617	801	ubiquitin-fold	hsa-miR-	−0.36	0.63	2009
			modifier 1	4458
ZNF644	NM_016620	802	zinc finger protein	hsa-miR-	−0.28	0.97	2005,
			644	4458			2007,
							2009
FZD3	NM_017412	803	frizzled family	hsa-miR-98	−0.2	>0.99
			receptor 3
RBM38	NM_017495	804	RNA binding motif	hsa-miR-	−0.25	0.98	2007,
			protein 38	4458			2009
STAB2	NM_017564	805	stabilin 2	hsa-miR-	−0.13	0.9	2005,
				4500			2007,
							2009
KIF21B	NM_017596	806	kinesin family	hsa-miR-	>−0.03	0.95	2007,
			member 21B	4458			2009
EIF2C4	NM_017629	807	eukaryotic translation	hsa-let-7d	−0.17	0.95	2005,
			initiation factor 2C, 4				2007,
							2009
BNC2	NM_017637	808	basonuclin 2	hsa-let-7g	−0.03	0.88	2005,
							2007,
							2009
KLHL24	NM_017644	809	kelch-like 24	hsa-miR-	>−0.01	0.87	2007,
			(Drosophila)	4458			2009
GDAP2	NM_017686	810	ganglioside induced	hsa-let-7d	−0.33	0.97	2009
			differentiation
			associated protein 2
FBXL12	NM_017703	811	F-box and leucine-	hsa-miR-	−0.32	0.92	2007,
			rich repeat protein 12	4458			2009
ANKRD49	NM_017704	812	ankyrin repeat	hsa-miR-	−0.25	0.9	2007,
			domain 49	4500			2009
UHRF1BP1	NM_017754	813	UHRF1 binding	hsa-miR-	−0.03	0.85
			protein 1	4500
INO80D	NM_017759	814	INO80 complex	hsa-miR-	−0.12	0.91	2005,
			subunit D	4500			2007,
							2009
CHD7	NM_017780	815	chromodomain	hsa-miR-	−0.07	0.86	2005,
			helicase DNA binding	4500			2007,
			protein 7				2009
SEMA4C	NM_017789	816	sema domain,	hsa-let-7i	−0.32	0.98	2005,
			immunoglobulin				2007,
			domain (Ig),				2009
			transmembrane
			domain (TM) and
			short cytoplasmic
			domain, (semaphorin)
			4C
CMTM6	NM_017801	817	CKLF-like MARVEL	hsa-let-7c	−0.16	<0.1
			transmembrane
			domain containing 6
HIF1AN	NM_017902	818	hypoxia inducible	hsa-miR-	−0.47	0.94	2009
			factor 1, alpha subunit	4500
			inhibitor
STX17	NM_017919	819	syntaxin 17	hsa-miR-	−0.16	0.97	2005,
				4500			2007,
							2009
USP47	NM_017944	820	ubiquitin specific	hsa-let-7d	−0.14	0.94	2007,
			peptidase 47				2009
PDPR	NM_017990	821	pyruvate	hsa-miR-	−0.3	0.95	2005,
			dehydrogenase	4500			2007,
			phosphatase				2009
			regulatory subunit
C9orf40	NM_017998	822	chromosome 9 open	hsa-let-7f	−0.55	0.76	2009
			reading frame 40
XKR8	NM_018053	823	XK, Kell blood group	hsa-miR-	−0.49	0.98	2007,
			complex subunit-	4458			2009
			related family,
			member 8
PRPF38B	NM_018061	824	PRP38 pre-mRNA	hsa-miR-	−0.32	0.26	2007,
			processing factor 38	4458			2009
			(yeast) domain
			containing B
IPO9	NM_018085	825	importin 9	hsa-miR-	−0.06	0.78	2009
				4500
FIGN	NM_018086	826	fidgetin	hsa-let-7d	−0.56	>0.99	2007,
							2009
CDCA8	NM_018101	827	cell division cycle	hsa-miR-	−0.17	0.94	2009
			associated 8	4458
FAM178A	NM_018121	828	family with sequence	hsa-miR-98	−0.24	0.98	2003,
			similarity 178,				2005,
			member A				2007,
							2009
LRRC20	NM_018205	829	leucine rich repeat	hsa-miR-	−0.04	0.87	2009
			containing 20	4458
ETNK2	NM_018208	830	ethanolamine kinase 2	hsa-miR-	−0.14	0.93	2005,
				4458			2007,
							2009
TMEM143	NM_018273	831	transmembrane	hsa-miR-	−0.23	0.9	2009
			protein 143	4500
BRF2	NM_018310	832	BRF2, subunit of	hsa-miR-	−0.43	<0.1	2009
			RNA polymerase III	4500
			transcription initiation
			factor, BRF1-like
DDX19A	NM_018332	833	DEAD (Asp-Glu-Ala-	hsa-miR-	−0.47	0.94	2007,
			As) box polypeptide	4500			2009
			19A
FGD6	NM_018351	834	FYVE, RhoGEF and	hsa-miR-	−0.32	0.99	2009
			PH domain	4458
			containing 6
ACER3	NM_018367	835	alkaline ceramidase 3	hsa-let-7d	−0.2	0.93
SYNJ2BP	NM_018373	836	synaptojanin 2	hsa-miR-	−0.11	0.85
			binding protein	4500
PAG1	NM_018440	837	phosphoprotein	hsa-miR-	−0.3	0.98	2009
			associated with	4458
			glycosphingolipid
			microdomains
1
ACTR10	NM_018477	838	actin-related protein	hsa-let-7f	−0.34	0.75	2009
			10 homolog (S. cerevisiae)
LGR4	NM_018490	839	leucine-rich repeat	hsa-miR-	−0.2	0.92	2005,
			containing G protein-	4500			2007,
			coupled receptor 4				2009
YOD1	NM_018566	840	YOD1 OTU	hsa-miR-	−0.57	>0.99	2007,
			deubiquinating	4500			2009
			enzyme 1 homolog
			(S. cerevisiae)
SLC16A10	NM_018593	841	solute carrier family	hsa-miR-	−0.14	0.86	2009
			16, member 10	4500
			(aromatic amino acid
			transporter)
ETNK1	NM_018638	842	ethanolamine kinase 1	hsa-miR-	−0.05	0.92	2007,
				4500			2009
B3GAT1	NM_018644	843	beta-1,3-	hsa-let-7d	−0.02	0.9	2009
			glucuronyltransferase 1
			(glucuronosyltransferase
			P)
BIN3	NM_018688	844	bridging integrator 3	hsa-let-7g	−0.22	0.92	2005,
							2007,
							2009
YIPF1	NM_018982	845	Yip1 domain family,	hsa-let-7d	−0.18	0.71
			member 1
SSH1	NM_018984	846	slingshot homolog 1	hsa-let-7a	−0.17	0.95	2005,
			(Drosophila)				2007,
							2009
SMCR7L	NM_019008	847	Smith-Magenis	hsa-let-7f	−0.08	0.94	2007,
			syndrome				2009
			chromosome region,
			candidate 7-like
CCDC93	NM_019044	848	coiled-coil domain	hsa-miR-	−0.07	0.62	2009
			containing 93	4458
CRCT1	NM_019060	849	cysteine-rich C-	hsa-miR-	−0.28	0.92	2009
			terminal 1	4500
CCDC76	NM_019083	850	coiled-coil domain	hsa-let-7f	−0.38	0.72
			containing 76
UBFD1	NM_019116	851	ubiquitin family	hsa-miR-	−0.1	0.87	2007,
			domain containing 1	4458			2009
TMEM234	NM_019118	852	transmembrane	hsa-let-7a	−0.37	0.92	2009
			protein 234
RNF20	NM_019592	853	ring finger protein 20	hsa-let-7g	−0.27	0.7	2005,
							2007
GPCPD1	NM_019593	854	glycerophosphocholine	hsa-miR-	−0.42	>0.99	2007,
			phosphodiesterase	4500			2009
			GDE1 homolog (S. cerevisiae)
ABCB9	NM_019624	855	ATP-binding cassette,	hsa-miR-98	−0.3	0.95	2005,
			sub-family B				2007,
			(MDR/TAP), member 9				2009
UGGT1	NM_020120	856	UDP-glucose	hsa-miR-	−0.22	0.98	2007,
			glycoprotein	4458			2009
			glucosyltransferase 1
KCMF1	NM_020122	857	potassium channel	hsa-let-7g	−0.02	0.93	2009
			modulatory factor 1
C1GALT1	NM_020156	858	core 1 synthase,	hsa-miR-98	−0.07	0.98
			glycoprotein-N-
			acetylgalactosamine
			3-beta-
			galactosyltransferase, 1
SLC12A9	NM_020246	859	solute carrier family	hsa-miR-	−0.29	0.98	2009
			12	4458
			(potassium/chloride
			transporters), member 9
MNT	NM_020310	860	MAX binding protein	hsa-let-7d	−0.02	0.93	2005,
							2007,
							2009
VANGL2	NM_020335	861	vang-like 2 (van	hsa-miR-	−0.06	>0.99	2007,
			gogh, Drosophila)	4458			2009
KIAA1244	NM_020340	862	KIAA1244	hsa-miR-	>−0.02	0.79
				4458
ENTPD7	NM_020354	863	ectonucleoside	hsa-let-7d	−0.17	0.94
			triphosphate
			diphosphohydrolase
7
AVEN	NM_020371	864	apoptosis, caspase	hsa-let-7i	−0.22	0.59
			activation inhibitor
SCYL3	NM_020423	865	SCY1-like 3 (S. cerevisiae)	hsa-let-7f	−0.18	0.93	2005,
							2007,
							2009
ASPHD2	NM_020437	866	aspartate beta-	hsa-miR-	−0.02	0.8
			hydroxylase domain	4458
			containing 2
GALNT1	NM_020474	867	UDP-N-acetyl-alpha-	hsa-miR-	−0.47	>0.99	2005,
			D-	4500			2007,
			galactosamine:polypeptide				2009
			N-
			acetylgalactosaminyltransferase 1
			(GalNAc-T1)
MRS2	NM_020662	868	MRS2 magnesium	hsa-let-7f	−0.48	0.91	2009
			homeostasis factor
			homolog (S. cerevisiae)
RAB22A	NM_020673	869	RAB22A, member	hsa-miR-	−0.15	0.88	2009
			RAS oncogene family	4500
ZNF512B	NM_020713	870	zinc finger protein	hsa-miR-	−0.31	>0.99	2007,
			512B	4458			2009
PLEKHH1	NM_020715	871	pleckstrin homology	hsa-miR-	−0.2	0.93	2007,
			domain containing,	4500			2009
			family H (with
			MyTH4 domain)
			member 1
NLN	NM_020726	872	neurolysin	hsa-miR-	−0.1	0.79
			(metallopeptidase M3	4458
			family)
INTS2	NM_020748	873	integrator complex	hsa-let-7a	−0.3	0.84	2007,
			subunit 2				2009
STARD9	NM_020759	874	StAR-related lipid	hsa-miR-	−0.7	0.79
			transfer (START)	4458
			domain containing 9
SRGAP1	NM_020762	875	SLIT-ROBO Rho	hsa-miR-	−0.08	0.73
			GTPase activating	4458
			protein 1
CASKIN1	NM_020764	876	CASK interacting	hsa-let-7i	−0.1	0.89	2005,
			protein 1				2007,
							2009
KCTD16	NM_020768	877	potassium channel	hsa-miR-	−0.19	0.87	2009
			tetramerisation	4458
			domain containing 16
RGAG1	NM_020769	878	retrotransposon gag	hsa-let-7f	−0.14	0.79	2005,
			domain containing 1				2007
MIB1	NM_020774	879	mindbomb homolog 1	hsa-let-7f	−0.15	>0.99	2007,
			(Drosophila)				2009
ALPK3	NM_020778	880	alpha-kinase 3	hsa-miR-	>−0.02	0.84	2009
				4458
PDP2	NM_020786	881	pyruvate	hsa-let-7b	−0.18	0.92	2009
			dehyrogenase
			phosphatase catalytic
			subunit
2
TAOK1	NM_020791	882	TAO kinase 1	hsa-let-7g	−0.07	0.91
ARHGAP20	NM_020809	883	Rho GTPase	hsa-let-7a	−0.11	0.93	2005,
			activating protein 20				2007,
							2009
KIAA1467	NM_020853	884	KIAA1467	hsa-miR-	−0.15	0.83	2009
				4458
ZSWIM5	NM_020883	885	zinc finger, SWIM-	hsa-miR-	−0.29	0.97	2009
			type containing 5	4458
PLXNA4	NM_020911	886	plexin A4	hsa-miR-	−0.15	>0.99	2009
				4500
SLC7A14	NM_020949	887	solute carrier family 7	hsa-let-7d	−0.07	0.94
			(orphan transporter),
			member 14
IGDCC4	NM_020962	888	immunoglobulin	hsa-let-7a	−0.24	0.98	2005,
			superfamily, DCC				2007,
			subclass, member 4				2009
SPTBN4	NM_020971	889	spectrin, beta, non-	hsa-miR-	−0.07	0.87	2007,
			erythrocytic 4	4458			2009
XK	NM_021083	890	X-linked Kx blood	hsa-miR-	−0.29	0.93	2009
			group (McLeod	4458
			syndrome)
MTMR3	NM_021090	891	myotubularin related	hsa-let-7a	−0.05	0.88	2009
			protein 3
SLC5A6	NM_021095	892	solute carrier family 5	hsa-let-7f	−0.21	0.93	2007,
			(sodium-dependent				2009
			vitamin transporter),
			member 6
COL14A1	NM_021110	893	collagen, type XIV,	hsa-miR-	−0.3	0.71	2007
			alpha 1	4500
PMAIP1	NM_021127	894	phorbol-12-myristate-	hsa-miR-	−0.17	0.9	2009
			13-acetate-induced	4500
			protein 1
FAM108C1	NM_021214	895	family with sequence	hsa-miR-98	−0.2	0.72
			similarity 108,
			member C1
RRAGD	NM_021244	896	Ras-related GTP	hsa-miR-	−0.1	0.93
			binding D	4500
CDH22	NM_021248	897	cadherin 22, type 2	hsa-miR-	−0.23	0.88
				4500
SNX6	NM_021249	898	sorting nexin 6	hsa-let-7c	−0.38	0.98	2009
SENP2	NM_021627	899	SUMO1/sentrin/SMT	hsa-miR-	−0.17	0.8	2005,
			3 specific peptidase 2	4458			2007,
							2009
TRIB2	NM_021643	900	tribbles homolog 2	hsa-miR-	−0.11	0.84	2005,
			(Drosophila)	4458			2007,
							2009
SPCS3	NM_021928	901	signal peptidase	hsa-miR-	−0.06	0.81
			complex subunit 3	4458
			homolog (S. cerevisiae)
FKBP10	NM_021939	902	FK506 binding	hsa-let-7d	−0.08	0.65
			protein 10, 65 kDa
TIA1	NM_022037	903	TIA1 cytotoxic	hsa-let-7f	−0.12	0.93
			granule-associated
			RNA binding protein
GAN	NM_022041	904	gigaxonin	hsa-miR-	−0.26	0.98	2005,
				4500			2007,
							2009
CERS2	NM_022075	905	ceramide synthase 2	hsa-let-7c	−0.11	0.69
PRSS22	NM_022119	906	protease, serine, 22	hsa-miR-98	−0.2	0.84
SNX16	NM_022133	907	sorting nexin 16	hsa-miR-	−0.23	0.94	2005,
				4500			2007,
							2009
XYLT1	NM_022166	908	xylosyltransferase I	hsa-miR-	>−0.03	0.99	2007,
				4458			2009
DNAJC1	NM_022365	909	DnaJ (Hsp40)	hsa-let-7d	−0.21	0.85	2005,
			homolog, subfamily				2007,
			C, member 1				2009
NSD1	NM_022455	910	nuclear receptor	hsa-miR-	−0.12	0.76
			binding SET domain	4458
			protein 1
HIF3A	NM_022462	911	hypoxia inducible	hsa-let-7b	−0.35	>0.99	2009
			factor 3, alpha subunit
ZMAT3	NM_022470	912	zinc finger, matrin-	hsa-miR-	>−0.01	0.73
			type 3	4458
TTC31	NM_022492	913	tetratricopeptide	hsa-miR-	−0.36	0.87	2009
			repeat domain 31	4458
MESDC1	NM_022566	914	mesoderm	hsa-miR-	−0.15	0.8	2005,
			development	4500			2007,
			candidate 1				2009
ELOVL4	NM_022726	915	ELOVL fatty acid	hsa-miR-	−0.16	0.94	2005,
			elongase 4	4500			2007,
							2009
FAM160B2	NM_022749	916	family with sequence	hsa-miR-	−0.06	0.92	2005,
			similarity 160,	4458			2007,
			member B2				2009
AEN	NM_022767	917	apoptosis enhancing	hsa-miR-	−0.31	0.93	2009
			nuclease	4458
RNF38	NM_022781	918	ring finger protein 38	hsa-let-7g	−0.13	0.7	2005,
							2007,
							2009
ANKRA2	NM_023039	919	ankyrin repeat, family	hsa-miR-	−0.19	0.84
			A (RFXANK-like), 2	4458
ZSWIM4	NM_023072	920	zinc finger, SWIM-	hsa-miR-	−0.03	0.79	2007
			type containing 4	4458
OTUB2	NM_023112	921	OTU domain,	hsa-miR-	−0.09	0.65
			ubiquitin aldehyde	4458
			binding 2
LRFN4	NM_024036	922	leucine rich repeat	hsa-miR-	−0.12	0.89	2007,
			and fibronectin type	4458			2009
			III domain containing 4
GNPTAB	NM_024312	923	N-acetylglucosamine-	hsa-miR-	−0.48	0.95	2007,
			1-phosphate	4500			2009
			transferase, alpha and
			beta subunits
EFHD2	NM_024329	924	EF-hand domain	hsa-let-7f	−0.19	0.97	2005,
			family, member D2				2007,
							2009
HOXD1	NM_024501	925	homeobox D1	hsa-miR-	−0.25	0.93	2005,
				4500			2007,
							2009
ADIPOR2	NM_024551	926	adiponectin receptor 2	hsa-let-7f	−0.16	0.94	2005,
							2007,
							2009
CCNJL	NM_024565	927	cyclin J-like	hsa-let-7f	−0.12	0.89	2009
SRD5A3	NM_024592	928	steroid 5 alpha-	hsa-let-7a	−0.4	0.84
			reductase 3
THAP9	NM_024672	929	THAP domain	hsa-miR-	−0.47	0.9
			containing 9	4458
LIN28A	NM_024674	930	lin-28 homolog A (C. elegans)	hsa-let-7i	−0.25	0.98	2005,
							2007,
							2009
KCTD17	NM_024681	931	potassium channel	hsa-miR-	−0.24	0.98	2007,
			tetramerisation	4458			2009
			domain containing 17
C15orf29	NM_024713	932	chromosome 15 open	hsa-miR-	−0.18	0.94	2005,
			reading frame 29	4458			2007,
							2009
MOBKL2B	NM_024761	933	MOB1, Mps One	hsa-miR-	>−0.03	0.93	2009
			Binder kinase	4458
			activator-like 2B
			(yeast)
ATP8B4	NM_024837	934	ATPase, class I, type	hsa-let-7a	−0.25	0.94	2009
			8B, member 4
EIF2C3	NM_024852	935	eukaryotic translation	hsa-let-7f	−0.13	0.83	2005,
			initiation factor 2C, 3				2007,
							2009
L2HGDH	NM_024884	936	L-2-hydroxyglutarate	hsa-let-7a	−0.05	0.98	2009
			dehydrogenase
C7orf58	NM_024913	937	chromosome 7 open	hsa-let-7g	−0.14	0.92	2005,
			reading frame 58				2007,
							2009
PHC3	NM_024947	938	polyhomeotic	hsa-let-7b	−0.05	0.84	2009
			homolog 3
			(Drosophila)
CEP135	NM_025009	939	centrosomal protein	hsa-let-7b	−0.34	0.95	2009
			135 kDa
ARHGEF15	NM_025014	940	Rho guanine	hsa-miR-	−0.23	0.94	2005,
			nucleotide exchange	4500			2007,
			factor (GEF) 15				2009
VCPIP1	NM_025054	941	valosin containing	hsa-miR-	>−0.01	0.76	2005,
			protein (p97)/p47	4458			2007
			complex interacting
			protein 1
FRAS1	NM_025074	942	Fraser syndrome 1	hsa-let-7b	−0.42	0.95	2005,
							2007,
							2009
NYNRIN	NM_025081	943	NYN domain and	hsa-let-7d	−0.32	>0.99	2007,
			retroviral integrase				2009
			containing
CHD9	NM_025134	944	chromodomain	hsa-let-7a	−0.07	0.74	2005,
			helicase DNA binding				2007
			protein 9
KIAA1539	NM_025182	945	KIAA1539	hsa-miR-	−0.25	0.92	2005,
				4458			2007,
							2009
EDEM3	NM_025191	946	ER degradation	hsa-miR-	−0.21	0.98	2007,
			enhancer,	4500			2009
			mannosidase alpha-
			like 3
TRIB1	NM_025195	947	tribbles homolog 1	hsa-miR-	−0.1	0.92	2005,
			(Drosophila)	4500			2007,
							2009
TRABD	NM_025204	948	TraB domain	hsa-miR-	−0.22	0.73	2007,
			containing	4458			2009
MED28	NM_025205	949	mediator complex	hsa-miR-	−0.45	0.32
			subunit 28	4500
SOST	NM_025237	950	sclerostin	hsa-miR-	−0.04	0.87	2009
				4500
LIMD2	NM_030576	951	LIM domain	hsa-let-7i	−0.38	0.95	2007,
			containing 2				2009
CPEB4	NM_030627	952	cytoplasmic	hsa-let-7d	−0.08	0.96	2005,
			polyadenylation				2007,
			element binding				2009
			protein 4
DUSP16	NM_030640	953	dual specificity	hsa-let-7d	−0.19	0.97	2005,
			phosphatase 16				2007,
							2009
C1orf21	NM_030806	954	chromosome 1 open	hsa-let-7g	−0.03	0.77
			reading frame 21
LBH	NM_030915	955	limb bud and heart	hsa-let-7g	−0.07	0.92	2005,
			development homolog				2007,
			(mouse)				2009
FAM103A1	NM_031452	956	family with sequence	hsa-miR-	−0.42	0.87	2009
			similarity 103,	4458
			member A1
SLC25A18	NM_031481	957	solute carrier family	hsa-miR-	−0.34	0.94	2005,
			25 (mitochondrial	4458			2007,
			carrier), member 18				2009
KCTD10	NM_031954	958	potassium channel	hsa-miR-	>−0.02	0.86	2009
			tetramerisation	4458
			domain containing 10
STARD3NL	NM_032016	959	STARD3 N-terminal	hsa-miR-	−0.15	0.91	2005,
			like	4458			2007,
							2009
STK40	NM_032017	960	serine/threonine	hsa-miR-	−0.25	0.95	2007,
			kinase 40	4458			2009
UTP15	NM_032175	961	UTP15, U3 small	hsa-let-7b	−0.16	0.79	2009
			nucleolar
			ribonucleoprotein,
			homolog (S. cerevisiae)
LOXL4	NM_032211	962	lysyl oxidase-like 4	hsa-let-7a	−0.13	0.94	2005,
							2007,
							2009
DDI2	NM_032341	963	DNA-damage	hsa-miR-	−0.46	0.98	2007,
			inducible 1 homolog	4500			2009
			2 (S. cerevisiae)
MEGF11	NM_032445	964	multiple EGF-like-	hsa-miR-	−0.05	0.88	2009
			domains 11	4500
DOT1L	NM_032482	965	DOT1-like, histone	hsa-miR-	N/A	0.99	2005,
			H3 methyltransferase	4458			2007,
			(S. cerevisiae)				2009
C6orf168	NM_032511	966	chromosome 6 open	hsa-miR-	−0.22	0.95	2009
			reading frame 168	4458
PARD6B	NM_032521	967	par-6 partitioning	hsa-let-7a	−0.29	0.95	2007,
			defective 6 homolog				2009
			beta (C. elegans)
USP38	NM_032557	968	ubiquitin specific	hsa-miR-	−0.33	0.85	2005,
			peptidase 38	4500			2007,
							2009
USP32	NM_032582	969	ubiquitin specific	hsa-let-7a	−0.2	0.93	2005,
			peptidase 32				2007,
							2009
LOXL3	NM_032603	970	lysyl oxidase-like 3	hsa-miR-	−0.1	0.79	2005,
				4458			2007,
							2009
FOXP1	NM_032682	971	forkhead box P1	hsa-let-7g	−0.04	0.85	2009
SFT2D3	NM_032740	972	SFT2 domain	hsa-let-7a	−0.34	<0.1	2009
			containing 3
LINGO1	NM_032808	973	leucine rich repeat	hsa-let-7d	−0.3	0.89	2009
			and Ig domain
			containing 1
ZNF341	NM_032819	974	zinc finger protein	hsa-let-7a	−0.42	<0.1
			341
PPP1R15B	NM_032833	975	protein phosphatase 1,	hsa-let-7c	−0.44	>0.99	2005,
			regulatory (inhibitor)				2007,
			subunit 15B				2009
CGNL1	NM_032866	976	cingulin-like 1	hsa-miR-	−0.2	0.94	2005,
				4458			2007,
							2009
RAB11FIP4	NM_032932	977	RAB11 family	hsa-let-7d	−0.27	>0.99	2003,
			interacting protein 4				2005,
			(class II)				2007,
							2009
C5orf62	NM_032947	978	chromosome 5 open	hsa-let-7d	−0.38	0.92	2007,
			reading frame 62				2009
ZCCHC3	NM_033089	979	zinc finger, CCHC	hsa-miR-	−0.16	0.93	2007,
			domain containing 3	4458			2009
NKD1	NM_033119	980	naked cuticle	hsa-let-7a	−0.2	0.95	2005,
			homolog 1				2007,
			(Drosophila)				2009
SCRT2	NM_033129	981	scratch homolog 2,	hsa-miR-	−0.05	0.88	2005,
			zinc finger protein	4458			2007,
			(Drosophila)				2009
SURF4	NM_033161	982	surfeit 4	hsa-let-7a	−0.02	0.87	2005,
							2007,
							2009
PURB	NM_033224	983	purine-rich element	hsa-miR-	−0.03	0.87	2009
			binding protein B	4458
RASL10B	NM_033315	984	RAS-like, family 10,	hsa-miR-	−0.06	0.94	2005,
			member B	4458			2007,
							2009
C20orf54	NM_033409	985	chromosome 20 open	hsa-miR-	−0.3	<0.1
			reading frame 54	4458
FAM125B	NM_033446	986	family with sequence	hsa-miR-	>−0.01	0.77	2007,
			similarity 125,	4458			2009
			member B
TSPYL5	NM_033512	987	TSPY-like 5	hsa-miR-	−0.24	0.58
				4458
TRIM41	NM_033549	988	tripartite motif	hsa-let-7f	−0.22	0.92
			containing 41
STARD13	NM_052851	989	StAR-related lipid	hsa-let-7g	−0.4	>0.99	2005,
			transfer (START)				2007,
			domain containing 13				2009
VPS26B	NM_052875	990	vacuolar protein	hsa-miR-	−0.15	0.9	2009
			sorting 26 homolog B	4500
			(S. pombe)
EGLN2	NM_053046	991	egl nine homolog 2	hsa-miR-	−0.16	0.93	2005,
			(C. elegans)	4500			2007,
							2009
CCND1	NM_053056	992	cyclin D1	hsa-let-7b	−0.12	0.99	2005,
							2007,
							2009
GALNTL2	NM_054110	993	UDP-N-acetyl-alpha-	hsa-miR-	−0.2	0.94	2005,
			D-	4500			2007,
			galactosamine:polypeptide				2009
			N-
			acetylgalactosaminyltransferase-
			like 2
C20orf112	NM_080616	994	chromosome 20 open	hsa-let-7b	−0.22	0.97
			reading frame 112
TMEM41A	NM_080652	995	transmembrane	hsa-miR-	−0.13	0.94
			protein 41A	4458
ADAMTS14	NM_080722	996	ADAM	hsa-miR-	−0.18	0.93	2007,
			metallopeptidase with	4458			2009
			thrombospondin type
			1 motif, 14
ZNF280B	NM_080764	997	zinc finger protein	hsa-let-7d	−0.34	0.98	2007,
			280B				2009
SOCS4	NM_080867	998	suppressor of	hsa-miR-	−0.15	0.91	2005,
			cytokine signaling 4	4500			2007,
							2009
KLHL6	NM_130446	999	kelch-like 6	hsa-let-7d	−0.32	0.95	2007,
			(Drosophila)				2009
TSPAN18	NM_130783	1000	tetraspanin 18	hsa-miR-	−0.08	0.94
				4500
UNC5A	NM_133369	1001	unc-5 homolog A (C. elegans)	hsa-let-7d	−0.12	0.9	2007,
							2009
GRIN3A	NM_133445	1002	glutamate receptor,	hsa-miR-	>−0.01	0.87	2009
			ionotropic, N-methyl-	4458
			D-aspartate 3A
PYGO2	NM_138300	1003	pygopus homolog 2	hsa-miR-	−0.07	0.92	2005,
			(Drosophila)	4500			2007,
							2009
DCAF15	NM_138353	1004	DDB1 and CUL4	hsa-miR-	−0.17	0.95	2007,
			associated factor 15	4458			2009
MARS2	NM_138395	1005	methionyl-tRNA	hsa-let-7d	−0.37	0.94	2009
			synthetase 2,
			mitochondrial
MARCH9	NM_138396	1006	membrane-associated	hsa-miR-	−0.09	0.78	2007,
			ring finger (C3HC4) 9	4500			2009
ZNF689	NM_138447	1007	zinc finger protein	hsa-miR-	−0.29	0.94	2009
			689	4500
CTHRC1	NM_138455	1008	collagen triple helix	hsa-miR-	−0.31	0.76	2009
			repeat containing 1	4458
C11orf84	NM_138471	1009	chromosome 11 open	hsa-miR-	−0.14	0.94	2005,
			reading frame 84	4458			2007,
							2009
H2AFV	NM_138635	1010	H2A histone family,	hsa-miR-	>−0.03	<0.1
			member V	4458
CD200R1	NM_138806	1011	CD200 receptor 1	hsa-let-7a	−0.38	0.82	2009
ADAMTS15	NM_139055	1012	ADAM	hsa-let-7i	−0.38	>0.99
			metallopeptidase with
			thrombospondin type
			1 motif, 15
LIPH	NM_139248	1013	lipase, member H	hsa-miR-	−0.29	0.94	2009
				4500
PPTC7	NM_139283	1014	PTC7 protein	hsa-let-7d	−0.06	0.85	2007,
			phosphatase homolog				2009
			(S. cerevisiae)
GNAT1	NM_144499	1015	guanine nucleotide	hsa-let-7g	−0.14	<0.1
			binding protein (G
			protein), alpha
			transducing activity
			polypeptide
1
CNOT6L	NM_144571	1016	CCR4-NOT	hsa-miR-	−0.02	0.92	2007,
			transcription	4458			2009
			complex, subunit 6-
			like
MAPK1IP1L	NM_144578	1017	mitogen-activated	hsa-miR-	−0.03	0.99	2009
			protein kinase 1	4458
			interacting protein 1-
			like
TEX261	NM_144582	1018	testis expressed 261	hsa-miR-	−0.04	0.8	2009
				4458
FOPNL	NM_144600	1019	FGFR1OP N-terminal	hsa-let-7a	−0.13	0.88	2009
			like
KLHL23	NM_144711	1020	kelch-like 23	hsa-let-7f	−0.2	0.95
			(Drosophila)
SMCR8	NM_144775	1021	Smith-Magenis	hsa-let-7d	−0.16	0.95	2009
			syndrome
			chromosome region,
			candidate 8
TSPEAR	NM_144991	1022	thrombospondin-type	hsa-let-7a	−0.16	0.9	2005,
			laminin G domain and				2007
			EAR repeats
TET3	NM_144993	1023	tet oncogene family	hsa-miR-98	−0.19	>0.99	2009
			member 3
CCNY	NM_145012	1024	cyclin Y	hsa-let-7d	−0.22	0.94
SLC2A12	NM_145176	1025	solute carrier family 2	hsa-miR-	−0.2	0.93	2007,
			(facilitated glucose	4500			2009
			transporter), member
			12
B3GNT7	NM_145236	1026	UDP-	hsa-miR-	−0.2	0.98
			GlcNAc:betaGal beta-	4458
			1,3-N-
			acetylglucosaminyltransferase 7
KIFC2	NM_145754	1027	kinesin family	hsa-miR-	−0.16	0.64
			member C2	4500
MTPN	NM_145808	1028	myotrophin	hsa-let-7d	−0.04	0.82	2005,
							2007,
							2009
SYT11	NM_152280	1029	synaptotagmin XI	hsa-miR-	−0.13	0.98	2005,
				4458			2007,
							2009
POGLUT1	NM_152305	1030	protein O-	hsa-let-7f	−0.14	0.87	2007,
			glucosyltransferase 1				2009
GRPEL2	NM_152407	1031	GrpE-like 2,	hsa-let-7i	−0.18	0.93	2009
			mitochondrial (E. coli)
RNF165	NM_152470	1032	ring finger protein	hsa-miR-	−0.19	>0.99	2007,
			165	4458			2009
ZNF362	NM_152493	1033	zinc finger protein	hsa-miR-	−0.08	0.98	2007,
			362	4458			2009
ARL6IP6	NM_152522	1034	ADP-ribosylation-like	hsa-miR-	−0.33	0.83
			factor 6 interacting	4500
			protein 6
SLC16A14	NM_152527	1035	solute carrier family	hsa-miR-98	−0.18	0.94	2007,
			16, member 14				2009
			(monocarboxylic acid
			transporter 14)
C7orf60	NM_152556	1036	chromosome 7 open	hsa-let-7a	−0.06	0.87	2005,
			reading frame 60				2007,
							2009
FAM116A	NM_152678	1037	family with sequence	hsa-miR-	−0.08	0.7
			similarity 116,	4458
			member A
SENP5	NM_152699	1038	SUMO1/sentrin	hsa-miR-	−0.17	0.94	2005,
			specific peptidase 5	4458			2007,
							2009
HOXA9	NM_152739	1039	homeobox A9	hsa-let-7d	−0.16	0.85	2005,
							2007,
							2009
SDK1	NM_152744	1040	sidekick homolog 1,	hsa-miR-	−0.14	0.95
			cell adhesion	4458
			molecule (chicken)
SCUBE3	NM_152753	1041	signal peptide, CUB	hsa-miR-	>−0.02	0.88	2005,
			domain, EGF-like 3	4458			2007,
							2009
RICTOR	NM_152756	1042	RPTOR independent	hsa-miR-	−0.14	0.98	2007,
			companion of MTOR,	4500			2009
			complex 2
YTHDF3	NM_152758	1043	YTH domain family,	hsa-miR-	−0.1	0.86	2005,
			member 3	4500			2007,
							2009
MFSD8	NM_152778	1044	major facilitator	hsa-miR-98	−0.33	0.91
			superfamily domain
			containing 8
COL24A1	NM_152890	1045	collagen, type XXIV,	hsa-let-7a	−0.19	0.92	2005,
			alpha 1				2007,
							2009
UHRF2	NM_152896	1046	ubiquitin-like with	hsa-let-7d	−0.41	0.89	2005,
			PHD and ring finger				2007,
			domains 2				2009
PXT1	NM_152990	1047	peroxisomal, testis	hsa-let-7f	−0.54	0.93	2009
			specific 1
NPHP3	NM_153240	1048	nephronophthisis 3	hsa-let-7f	−0.49	>0.99	2009
			(adolescent)
BRWD3	NM_153252	1049	bromodomain and	hsa-let-7i	−0.12	0.95
			WD repeat domain
			containing 3
ATXN7L2	NM_153340	1050	ataxia 7-like 2	hsa-miR-	−0.18	0.75	2007
				4458
GPR26	NM_153442	1051	G protein-coupled	hsa-miR-	−0.17	0.95	2009
			receptor 26	4500
LCORL	NM_153686	1052	ligand dependent	hsa-miR-	−0.2	0.78	2007,
			nuclear receptor	4458			2009
			corepressor-like
FAM43A	NM_153690	1053	family with sequence	hsa-miR-	−0.16	0.82	2009
			similarity 43, member A	4458
TTL	NM_153712	1054	tubulin tyrosine ligase	hsa-miR-	−0.08	0.94	2007,
				4458			2009
ATP2A2	NM_170665	1055	ATPase, Ca++	hsa-miR-	−0.28	0.96	2005,
			transporting, cardiac	4500			2007
			muscle, slow twitch 2
IL28RA	NM_170743	1056	interleukin 28	hsa-miR-	−0.17	0.72
			receptor, alpha	4458
			(interferon, lambda
			receptor)
RDH10	NM_172037	1057	retinol dehydrogenase	hsa-miR-98	−0.19	0.95	2005,
			10 (all-trans)				2007,
							2009
DCUN1D3	NM_173475	1058	DCN1, defective in	hsa-miR-	−0.21	0.98	2007,
			cullin neddylation 1,	4500			2009
			domain containing 3
			(S. cerevisiae)
LSM11	NM_173491	1059	LSM11, U7 small	hsa-let-7b	−0.35	0.94	2005,
			nuclear RNA				2007,
			associated				2009
SLC38A9	NM_173514	1060	solute carrier family	hsa-miR-	−0.17	0.82	2005,
			38, member 9	4458			2007
KLHDC8B	NM_173546	1061	kelch domain	hsa-miR-	−0.35	0.98	2009
			containing 8B	4458
RFX6	NM_173560	1062	regulatory factor X, 6	hsa-let-7d	−0.24	0.95	2005,
							2007,
							2009
PRR14L	NM_173566	1063	proline rich 14-like	hsa-miR-	−0.03	0.9
				4458
UBN2	NM_173569	1064	ubinuclein 2	hsa-miR-	>−0.02	0.94	2009
				4458
PGM2L1	NM_173582	1065	phosphoglucomutase	hsa-miR-	−0.05	>0.99	2005,
			2-like 1	4458			2007,
							2009
ANKRD52	NM_173595	1066	ankyrin repeat	hsa-miR-	>−0.06	0.96	2009
			domain 52	4458
RIMKLA	NM_173642	1067	ribosomal	hsa-let-7b	−0.08	<0.1
			modification protein
			rimK-like family
			member A
CCDC141	NM_173648	1068	coiled-coil domain	hsa-miR-	−0.17	0.95
			containing 141	4458
SLC9A9	NM_173653	1069	solute carrier family 9	hsa-miR-	−0.11	0.89	2005,
			(sodium/hydrogen	4458			2007,
			exchanger), member 9				2009
C3orf64	NM_173654	1070	chromosome 3 open	hsa-let-7d	−0.21	>0.99	2007,
			reading frame 64				2009
CRB2	NM_173689	1071	crumbs homolog 2	hsa-miR-	−0.12	0.93	2009
			(Drosophila)	4458
PRTG	NM_173814	1072	protogenin	hsa-miR-98	−0.56	>0.99	2007,
							2009
SREK1IP1	NM_173829	1073	SREK1-interacting	hsa-let-7g	−0.12	0.95	2007,
			protein 1				2009
FAM84B	NM_174911	1074	family with sequence	hsa-miR-	−0.13	0.58	2007
			similarity 84, member B	4500
ZPLD1	NM_175056	1075	zona pellucida-like	hsa-miR-	−0.2	0.94	2009
			domain containing 1	4458
TXLNA	NM_175852	1076	taxilin alpha	hsa-let-7b	−0.1	0.9	2009
CHSY3	NM_175856	1077	chondroitin sulfate	hsa-miR-	−0.17	0.75	2007
			synthase 3	4500
C19orf39	NM_175871	1078	chromosome 19 open	hsa-let-7d	−0.32	0.77
			reading frame 39
ANKRD43	NM_175873	1079	ankyrin repeat	hsa-miR-	−0.22	0.94	2007,
			domain 43	4500			2009
FLJ36031	NM_175884	1080	hypothetical protein	hsa-let-7g	−0.31	0.88	2007,
			FLJ36031				2009
C5orf51	NM_175921	1081	chromosome 5 open	hsa-miR-	−0.26	0.94	2005,
			reading frame 51	4458			2007,
							2009
PRR18	NM_175922	1082	proline rich 18	hsa-miR-	−0.13	0.72
				4500
CXorf36	NM_176819	1083	chromosome X open	hsa-let-7e	−0.02	0.81
			reading frame 36
PDE12	NM_177966	1084	phosphodiesterase 12	hsa-let-7a	−0.25	0.98	2007,
							2009
SESTD1	NM_178123	1085	SEC14 and spectrin	hsa-miR-	−0.19	0.92
			domains 1	4500
SNAI3	NM_178310	1086	snail homolog 3	hsa-let-7i	−0.18	0.56	2007
			(Drosophila)
TMEM26	NM_178505	1087	transmembrane	hsa-miR-	−0.1	0.93	2009
			protein 26	4458
NAT8L	NM_178557	1088	N-acetyltransferase 8-	hsa-miR-98	−0.05	0.94
			like (GCN5-related,
			putative)
RSPO2	NM_178565	1089	R-spondin 2	hsa-let-7d	−0.3	0.88	2007,
							2009
MTDH	NM_178812	1090	metadherin	hsa-miR-98	−0.1	0.95
AGPAT6	NM_178819	1091	1-acylglycerol-3-	hsa-let-7d	−0.24	0.98
			phosphate O-
			acyltransferase 6
			(lysophosphatidic
			acid acyltransferase,
			zeta)
MFSD4	NM_181644	1092	major facilitator	hsa-let-7a	−0.31	0.98
			superfamily domain
			containing 4
TMTC3	NM_181783	1093	transmembrane and	hsa-miR-	−0.01	0.79
			tetratricopeptide	4500
			repeat containing 3
SLC46A3	NM_181785	1094	solute carrier family	hsa-miR-	−0.13	0.74
			46, member 3	4500
USP12	NM_182488	1095	ubiquitin specific	hsa-let-7f	−0.37	0.9
			peptidase 12
DDX26B	NM_182540	1096	DEAD/H (Asp-Glu-	hsa-let-7f	−0.25	0.94	2009
			Ala-Asp/His) box
			polypeptide 26B
GDPD1	NM_182569	1097	glycerophosphodiester	hsa-miR-	−0.19	0.94	2005,
			phosphodiesterase	4458			2007,
			domain containing 1				2009
SP8	NM_182700	1098	Sp8 transcription	hsa-miR-	−0.16	0.92	2007,
			factor	4458			2009
ARRDC4	NM_183376	1099	arrestin domain	hsa-miR-	−0.15	0.94	2005,
			containing 4	4458			2007,
							2009
KIF1B	NM_183416	1100	kinesin family	hsa-miR-	−0.07	0.88
			member 1B	4458
TMEM65	NM_194291	1101	transmembrane	hsa-miR-	−0.2	0.94	2007,
			protein 65	4500			2009
SLC16A9	NM_194298	1102	solute carrier family	hsa-miR-	−0.25	0.9	2009
			16, member 9	4458
			(monocarboxylic acid
			transporter 9)
E2F6	NM_198256	1103	E2F transcription	hsa-let-7c	−0.21	0.98	2007,
			factor 6				2009
ANKRD46	NM_198401	1104	ankyrin repeat	hsa-miR-	−0.19	0.87	2009
			domain 46	4458
NRK	NM_198465	1105	Nik related kinase	hsa-let-7i	−0.07	0.94	2005,
							2007,
							2009
NHLRC2	NM_198514	1106	NHL repeat	hsa-let-7d	−0.08	0.83
			containing 2
ZNF710	NM_198526	1107	zinc finger protein	hsa-let-7d	−0.24	>0.99	2003,
			710				2007,
							2009
TMEM110	NM_198563	1108	transmembrane	hsa-miR-	−0.24	0.94	2007,
			protein 110	4458			2009
RAB15	NM_198686	1109	RAB15, member	hsa-miR-	−0.16	0.94	2005,
			RAS onocogene	4458			2007,
			family				2009
DHX57	NM_198963	1110	DEAH (Asp-Glu-Ala-	hsa-miR-	−0.25	0.88	2005,
			Asp/His) box	4500			2007,
			polypeptide 57				2009
MED8	NM_201542	1111	mediator complex	hsa-let-7f	−0.42	0.92
			subunit 8
ZNF784	NM_203374	1112	zinc finger protein	hsa-miR-	−0.21	0.83	2009
			784	4458
PPAPDC2	NM_203453	1113	phosphatidic acid	hsa-miR-	−0.2	0.94	2007,
			phosphatase type 2	4500			2009
			domain containing 2
SPRYD4	NM_207344	1114	SPRY domain	hsa-miR-	−0.2	0.94	2007,
			containing 4	4458			2009
FREM2	NM_207361	1115	FRAS1 related	hsa-miR-	−0.07	0.94	2009
			extracellular matrix	4458
			protein 2
C10orf140	NM_207371	1116	chromosome 10 open	hsa-let-7d	−0.13	0.85	2005,
			reading frame 140				2007,
							2009

TABLE S3

3′ UTR sequences.

	Human FNTB
	(SEQ ID NO: 1117)
	5′AGGACCTGGGTCCCGGCAGCTCTTTGCTCACCCATCTCCCCAG

	TCAGACAAGGTTTATACGTTTCAATACATACTGCATTCTGTGCTA

	CACAAGCCTTAGCCTCAGTGGAGCTGTGGTTCTCTTGGTACTTTC

	TTGTCAAACAAAACCAATGGCTCTGGGTTTGGAGAACACAGTGGC

	TGGTTTTAAAAATTCTTTCCACACCTGTCAAACCAAAAATCTATC

	AGCCCACGTGGTGTGGTTGGTGAACAGTGCATGCCAGGAGGAAGC

	AGTCCCTCCTCACCAGCTCTCCAGCCAGGACGATCACACAGAGAT

	GAATGGCATCTGAGTATTACGGCATCCAGAGCCACTGCTGACTCC

	CACTTGCACGCCACCATTCAGTCACCAGACTGGGTGCCCTCCGAT

	GGGTGGAATAAGTCTGCTTCATGCCAAGGCTGGGCTTTGGGTCCC

	ACCAAGATGAGTTCTCTGTAAGACTGTGGTGGAGTTGCACCAGGA

	GGTGCCTCTGCCTCTCGACTTGCACCCTGGTCATTTGTAAGGGAA

	AAGAGCTGGAGGTGGGGAGAGAAAGATCTCCTTCAGTTGGGAGTC

	CTTCCACTTCAACACTGGAGAACTGAGCCTTGCATCTCTCCAGGG

	TCCAAGGCCACGCTTGGTGCACAGGCAAGACTTGCTTCAGCCCCA

	GGTGTGGTGACTTAGACCTAGGAAACCAATTATGAGTGGAAAGTG

	ACCCTCTAGTTCAACTGTGCCAGAGGAAACAGCCCTCCAGTGCCC

	ACCTGCCTCACTCCTCCCTATCATGTACCGTGAAAACCCCCTCTG

	ATGGCCTCAAGGCAGTGCCTGCAGGCCGAGGCCCTTCTGGGGGTT

	TCTATCTTTCTTCCACCAGACTCCAAGCCCACTCTCCTCCAAGAC

	TGTGTTGTCTTTTCTCACCAAGAGTATTAACACTACTAAGTCTTT

	CACCTTAACTTATGACTCAGGATTTATTCACGTCCTGCCCACTCT

	AGGCTCACAGGAATAAAATCAAGTGCTAGACACACTGGCTGCTAC

	TAAGGCACTAGCCTCTGTAGCTGGTGGTGGCAGCGTGGGGTGCCG

	CCCAGCGTGCTGGGTCCTGGCAGTGCCTCTGCTGTGCTGCACATT

	GAGCCCTTTCTCAGTCAGTGGAGTATCAAGTTGGGCCATCTGTCT

	ACTGACCTGGCCTTCATGTAAGCAGCTGTGGGCTGCGGGCAGACA

	GGAGCTCAGAGATGCAGCATGAGGCGCTTAGAAAAACCTGGCCAT

	TTGCTGCCTCTAATTCCCTTTTGCTTTG-3′

	Zebrafish Fntb
	(SEQ ID NO: 1118)
	5′ACTATCTATTTTTCAACTGTAAACATATTGGGGGATTTGGGCT

	AGCTTGAACTGTGCAGAGGAATCTTCTGTAAATGCTGTAGCCAAA

	TGTCCTTTGCTGGTGTGTACAGCGTCTACAGATGGAGAAACACTC

	TGAGGCCTGATCTGCTGCTGCTTCAGCATGAGGTTATGGGTTTTA

	GCCAGGATAAACAGTAGAGACTGACTAGGTAAGGTGTTTTTTCAT

	GCACCAGTCATTGATTGATATTTTATTGCACATGCCAGCTTATTT

	TGGGCAAAATGTTCCCAGAGTGTACGTGGCAGTGGGTTCTGGACA

	GAAATAAAGACTTGGATGGAAA-3′

	Zebrafish Smarca5
	(SEQ ID NO: 1119)
	5′GCAGGCAGGCATTTCACACACCTCACTCGGCGAGGAGCTTCAG

	TACAGCAATACTGCATTGATTGTTACGGGTCCCACTCATGTACTG

	TATGGATTTGCAGCACTGATCCTCGCCTCTCAAGTAGCTTGGCCT

	TCTTAACAAGGTGTAGAGTTGTAAATTAGGTCTCTTTTAGTTATA

	TAATGTAACTACGGCTGTGCTGTCGGATGTGTTTTGTATTTATGG

	CTACTTCAAATTTTTTTTTGTACCACATTCCATTTGCTTGTATCA

	GTTTAATTTGCAGTCTTTACCCCCTCATTATTAGTGTCTTCAGTA

	TTGTATTGTCTCTGTATCCGCCATTGGAAAGTGACTAATAAATGT

	GGTTTTTATAAATGCTGCTCTGTATGTTTCCTACAAATAAATGTA

	ATGTCTTTTGCCTTGTA-3′

TABLE S4

MiR mimics and antagomiR sequences.

	miRs used	Sequence

	Dre-miR-100 mimic	5′-AACCCGUAGAUCCG
		AACUUGUG-3′
		(SEQ ID NO: 1120)
		5′-CAAGCUUGUAUCUA
		UAGGUAUC-3′
		(SEQ ID NO: 1121)

	Dre-miR-99 mimic	5′-AACCCGUAGAUCCG
		AUCUUGUG-3′
		(SEQ ID NO: 1122)
		5′-CAAGCUCGAUUCUA
		UGGGUCUC-3′
		(SEQ ID NO: 1123)

	Dre-miR-99 inhibitor	5′-ACAAGTTCGGATCT
		ACGGGT-3′
		(SEQ ID NO: 1124)

	Dre-miR-100 inhibitor	5′-ACAAGATCGGATCT
		ACGGGT-3′
		(SEQ ID NO: 1125)

	GFP siRNA	5′-GGCUACGUCCAGGA
	(delivery control)	GCGCACC-3′
		(SEQ ID NO: 1126)
		5′-GGUGCGCUCCUGGA
		CGUAGCC-3′-Cy5
		(SEQ ID NO: 1127)

	Fntb morpholino	5′-GCGCCTCTTCCATG
	(translation-blocking)	ATGAGCTCTCA-3′
		(SEQ ID NO: 1128)

	Smarca5 morpholino	5′-CTTCTTCCCGCTGC
	(translation-blocking)	TGCTCCATGCT-3′
		(SEQ ID NO: 1129)

Claims

What is claimed is:

1. A method of treating a subject with heart damage, comprising:

administering to the subject a therapeutically effective amount of a nucleic acid encoding an antagonist of a micro RNA (miR) 99 micro RNA, a nucleic acid encoding an antagonist of a miR 100 micro RNA, a nucleic acid encoding an antagonist of a let-7a micro RNA, and a nucleic acid encoding an antagonist of a let-7c micro RNA,

thereby inducing proliferation of cardiomyocytes and treating the heart damage in the subject.

2. The method of claim 1, comprising administering to the subject a therapeutically effective amount of

a) a lentiviral vector or an adeno-associated viral (AAV) vector comprising the nucleic acid encoding the antagonist of the miR 99 micro RNA and the nucleic acid encoding the antagonist of the miR 100 micro RNA, and

b) a lentiviral vector or an AAV vector encoding the nucleic acid encoding the antagonist of the let-7a micro RNA and the antagonist of the let-7c micro RNA.

3. The method of claim 2, wherein a) the lentiviral vector or the AAV vector comprising the nucleic acid encoding the antagonist of the miR 99 micro RNA and the antagonist of the miR 100 micro RNA, and b) the lentiviral vector or the AAV vector encoding the nucleic acid encoding the antagonist of the let-7a micro RNA and the antagonist of the let-7c micro RNA, are the same vector.

4. The method of claim 2, wherein a) the lentiviral vector or the AAV vector comprising the nucleic acid encoding the antagonist of the miR 99 micro RNA and the antagonist of the miR 100 micro RNA, and b) the lentiviral vector or the AAV vector encoding the nucleic acid encoding the antagonist of the let-7a, are different vectors.

5. The method of claim 1, comprising administering to the subject a therapeutically effective amount of a nucleic acid molecule comprising the nucleic acid sequence as set forth in SEQ ID NO:1124 or SEQ ID NO:1125.

6. The method of claim 5, comprising administering to the subject the therapeutically effective amount of the nucleic acid molecule comprising the nucleic acid sequence set forth as SEQ ID NO: 1124.

7. The method of claim 5, comprising administering to the subject the therapeutically effective amount of the nucleic acid molecule comprising the nucleic acid sequence set forth as SEQ ID NO: 1125.

8. The method of claim 1, further comprising measuring the proliferation of cardiomyocytes in the subject.

9. The method of claim 1, wherein the method regenerates cardiac tissue in the subject.

10. The method of claim 1, wherein the subject is a mammal.

11. The method of claim 10, wherein the mammal is a human.

12. The method of claim 10, wherein the method improves ejection fraction in the subject.

13. The method of claim 1, wherein the heart damage is a myocardial infarction.

14. The method of claim 13, wherein the method reduces fibrotic scarring and/or infarct size in the subject.

15. A method of regenerating heart muscle in a subject, comprising

administering to the subject a therapeutically effective amount of a small molecule that modulates expression or activity of a miR 99 micro RNA-regulated protein, a small molecule that modulates expression or activity of a miR 100 micro RNA-regulated protein, a small molecule that modulates expression or activity of a let-7a micro RNA -regulated protein, and small molecule that modulates expression or activity of a let-7c micro RNA regulated protein, thereby forming a treated cardiomyocyte,

thereby inducing proliferation of cardiomyocytes and regenerating heart muscle in the subject.

16. The method of claim 15, wherein the small molecule that modulates expression or activity of the miR 99 micro RNA-regulated protein, the small molecule that modulates expression or activity of the miR 100 micro RNA-regulated protein, the small molecule that modulates expression or activity of the let-7a micro RNA -regulated protein, and/or the small molecule that modulates expression or activity of the let-7c micro RNA regulated protein is a synthetic micro RNA molecule.

17. The method of claim 15, further comprising measuring the proliferation of cardiomyocytes in the subject.

18. The method of claim 15, wherein the subject is human.

19. The method of claim 15, wherein the subject is a mammal.

20. The method of claim 15, wherein the mammal is a human.

21. The method of claim 15, wherein the method improves ejection fraction in the subject.

22. The method of claim 15, wherein the subject has a myocardial infarction.

23. The method of claim 22, wherein the method reduces fibrotic scarring and/or infarct size in the subject.