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Identification and characterization of FGF2-dependent mRNA: microRNA networks during lens fiber cell differentiation.

Wolf L, Gao CS, Gueta K, Xie Q, Chevallier T, Podduturi NR, Sun J, Conte I, Zelenka PS, Ashery-Padan R, Zavadil J, Cvekl A - G3 (Bethesda) (2013)

Bottom Line: Specific miRNA:mRNA interaction networks were predicted for c-Maf, N-Myc, and Nfib (DNA-binding transcription factors); Cnot6, Cpsf6, Dicer1, and Tnrc6b (RNA to miRNA processing); and Ash1l, Med1/PBP, and Kdm5b/Jarid1b/Plu1 (chromatin remodeling).Three miRNAs, including miR-143, miR-155, and miR-301a, down-regulated expression of c-Maf in the 3'-UTR luciferase reporter assays.These present studies demonstrate for the first time global impact of activated FGF signaling in lens cell culture system and predicted novel gene regulatory networks connected by multiple miRNAs that regulate lens differentiation.

View Article: PubMed Central - PubMed

Affiliation: Department of Genetics, Albert Einstein College of Medicine, Bronx, New York 10461.

ABSTRACT
MicroRNAs (miRNAs) and fibroblast growth factor (FGF) signaling regulate a wide range of cellular functions, including cell specification, proliferation, migration, differentiation, and survival. In lens, both these systems control lens fiber cell differentiation; however, a possible link between these processes remains to be examined. Herein, the functional requirement for miRNAs in differentiating lens fiber cells was demonstrated via conditional inactivation of Dicer1 in mouse (Mus musculus) lens. To dissect the miRNA-dependent pathways during lens differentiation, we used a rat (Rattus norvegicus) lens epithelial explant system, induced by FGF2 to differentiate, followed by mRNA and miRNA expression profiling. Transcriptome and miRNome analysis identified extensive FGF2-regulated cellular responses that were both independent and dependent on miRNAs. We identified 131 FGF2-regulated miRNAs. Seventy-six of these miRNAs had at least two in silico predicted and inversely regulated target mRNAs. Genes modulated by the greatest number of FGF-regulated miRNAs include DNA-binding transcription factors Nfib, Nfat5/OREBP, c-Maf, Ets1, and N-Myc. Activated FGF signaling influenced bone morphogenetic factor/transforming growth factor-β, Notch, and Wnt signaling cascades implicated earlier in lens differentiation. Specific miRNA:mRNA interaction networks were predicted for c-Maf, N-Myc, and Nfib (DNA-binding transcription factors); Cnot6, Cpsf6, Dicer1, and Tnrc6b (RNA to miRNA processing); and Ash1l, Med1/PBP, and Kdm5b/Jarid1b/Plu1 (chromatin remodeling). Three miRNAs, including miR-143, miR-155, and miR-301a, down-regulated expression of c-Maf in the 3'-UTR luciferase reporter assays. These present studies demonstrate for the first time global impact of activated FGF signaling in lens cell culture system and predicted novel gene regulatory networks connected by multiple miRNAs that regulate lens differentiation.

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A summary analysis of miRNA connectivity and their FGF2-mediated inducibility. (A) Top 10 ranking of FGF2-regulated “early” miRNAs depending on the number of their predicted/regulated target genes. (B) Top 10 ranking of FGF2-regulated “late” miRNAs depending on the number of their predicted/regulated target genes. The complete lists of genes/miRNAs are shown in File S3, A and B.
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fig5: A summary analysis of miRNA connectivity and their FGF2-mediated inducibility. (A) Top 10 ranking of FGF2-regulated “early” miRNAs depending on the number of their predicted/regulated target genes. (B) Top 10 ranking of FGF2-regulated “late” miRNAs depending on the number of their predicted/regulated target genes. The complete lists of genes/miRNAs are shown in File S3, A and B.

Mentions: It has been shown earlier that a single miRNA can recognize hundreds of target mRNAs and that multiple miRNAs can regulate expression of genes with joint roles in a specific pathway and connect multiple genes into highly complex regulatory hubs (Ivey and Srivastava 2010; Pauli et al. 2011). To address miRNA connectivity in the present study, we used human TargetScan in combination with ingenuity pathway analysis because this platform allows identification of inversely correlated miRNA and mRNA patterns in a batch mode with adjustable prediction score level. Using the starting numbers of 66 “early” and 65 “late” miRNAs, we found that this procedure mapped 31 and 45 miRNAs, with 549 and 531 target mRNAs, respectively. In sum, these 76 miRNAs regulated 1080 mRNAs. The top-ten most connected miRNAs, divided in “early” and “late” sets, are shown in Figure 5, A and B, respectively. The total number of their target genes is between 114 (1st rank, “early” miR-495) and 45 (10th rank, “early” miR-31 and miR-133b) connections. Notably, a number of genes appeared to be regulated by more than ten distinct miRNAs. For example, Cpsf6 and Tnrc6b (“early” group) are regulated by 11 distinct miRNAs. In the “late” group, Aak1, Cnot6, and Nfat5 were regulated by 16, 11, and 13 different FGF2-dependent miRNAs, respectively. From this analysis, we concluded that the present experimental set is enriched for specific mRNAs that are coregulated by a large number of distinct FGF2-regulated miRNAs, raising the possibility that these novel FGF2/miRNA-dependent gene regulatory networks (GRNs) play major roles during lens fiber cell differentiation.


Identification and characterization of FGF2-dependent mRNA: microRNA networks during lens fiber cell differentiation.

Wolf L, Gao CS, Gueta K, Xie Q, Chevallier T, Podduturi NR, Sun J, Conte I, Zelenka PS, Ashery-Padan R, Zavadil J, Cvekl A - G3 (Bethesda) (2013)

A summary analysis of miRNA connectivity and their FGF2-mediated inducibility. (A) Top 10 ranking of FGF2-regulated “early” miRNAs depending on the number of their predicted/regulated target genes. (B) Top 10 ranking of FGF2-regulated “late” miRNAs depending on the number of their predicted/regulated target genes. The complete lists of genes/miRNAs are shown in File S3, A and B.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3852386&req=5

fig5: A summary analysis of miRNA connectivity and their FGF2-mediated inducibility. (A) Top 10 ranking of FGF2-regulated “early” miRNAs depending on the number of their predicted/regulated target genes. (B) Top 10 ranking of FGF2-regulated “late” miRNAs depending on the number of their predicted/regulated target genes. The complete lists of genes/miRNAs are shown in File S3, A and B.
Mentions: It has been shown earlier that a single miRNA can recognize hundreds of target mRNAs and that multiple miRNAs can regulate expression of genes with joint roles in a specific pathway and connect multiple genes into highly complex regulatory hubs (Ivey and Srivastava 2010; Pauli et al. 2011). To address miRNA connectivity in the present study, we used human TargetScan in combination with ingenuity pathway analysis because this platform allows identification of inversely correlated miRNA and mRNA patterns in a batch mode with adjustable prediction score level. Using the starting numbers of 66 “early” and 65 “late” miRNAs, we found that this procedure mapped 31 and 45 miRNAs, with 549 and 531 target mRNAs, respectively. In sum, these 76 miRNAs regulated 1080 mRNAs. The top-ten most connected miRNAs, divided in “early” and “late” sets, are shown in Figure 5, A and B, respectively. The total number of their target genes is between 114 (1st rank, “early” miR-495) and 45 (10th rank, “early” miR-31 and miR-133b) connections. Notably, a number of genes appeared to be regulated by more than ten distinct miRNAs. For example, Cpsf6 and Tnrc6b (“early” group) are regulated by 11 distinct miRNAs. In the “late” group, Aak1, Cnot6, and Nfat5 were regulated by 16, 11, and 13 different FGF2-dependent miRNAs, respectively. From this analysis, we concluded that the present experimental set is enriched for specific mRNAs that are coregulated by a large number of distinct FGF2-regulated miRNAs, raising the possibility that these novel FGF2/miRNA-dependent gene regulatory networks (GRNs) play major roles during lens fiber cell differentiation.

Bottom Line: Specific miRNA:mRNA interaction networks were predicted for c-Maf, N-Myc, and Nfib (DNA-binding transcription factors); Cnot6, Cpsf6, Dicer1, and Tnrc6b (RNA to miRNA processing); and Ash1l, Med1/PBP, and Kdm5b/Jarid1b/Plu1 (chromatin remodeling).Three miRNAs, including miR-143, miR-155, and miR-301a, down-regulated expression of c-Maf in the 3'-UTR luciferase reporter assays.These present studies demonstrate for the first time global impact of activated FGF signaling in lens cell culture system and predicted novel gene regulatory networks connected by multiple miRNAs that regulate lens differentiation.

View Article: PubMed Central - PubMed

Affiliation: Department of Genetics, Albert Einstein College of Medicine, Bronx, New York 10461.

ABSTRACT
MicroRNAs (miRNAs) and fibroblast growth factor (FGF) signaling regulate a wide range of cellular functions, including cell specification, proliferation, migration, differentiation, and survival. In lens, both these systems control lens fiber cell differentiation; however, a possible link between these processes remains to be examined. Herein, the functional requirement for miRNAs in differentiating lens fiber cells was demonstrated via conditional inactivation of Dicer1 in mouse (Mus musculus) lens. To dissect the miRNA-dependent pathways during lens differentiation, we used a rat (Rattus norvegicus) lens epithelial explant system, induced by FGF2 to differentiate, followed by mRNA and miRNA expression profiling. Transcriptome and miRNome analysis identified extensive FGF2-regulated cellular responses that were both independent and dependent on miRNAs. We identified 131 FGF2-regulated miRNAs. Seventy-six of these miRNAs had at least two in silico predicted and inversely regulated target mRNAs. Genes modulated by the greatest number of FGF-regulated miRNAs include DNA-binding transcription factors Nfib, Nfat5/OREBP, c-Maf, Ets1, and N-Myc. Activated FGF signaling influenced bone morphogenetic factor/transforming growth factor-β, Notch, and Wnt signaling cascades implicated earlier in lens differentiation. Specific miRNA:mRNA interaction networks were predicted for c-Maf, N-Myc, and Nfib (DNA-binding transcription factors); Cnot6, Cpsf6, Dicer1, and Tnrc6b (RNA to miRNA processing); and Ash1l, Med1/PBP, and Kdm5b/Jarid1b/Plu1 (chromatin remodeling). Three miRNAs, including miR-143, miR-155, and miR-301a, down-regulated expression of c-Maf in the 3'-UTR luciferase reporter assays. These present studies demonstrate for the first time global impact of activated FGF signaling in lens cell culture system and predicted novel gene regulatory networks connected by multiple miRNAs that regulate lens differentiation.

Show MeSH
Related in: MedlinePlus