• Cdk9 Regulates cell growth, differentiation, and apoptosis of Myoblast Cells by Modulation of myomiRs Expression
  • Vahideh Tarhriz,1,* leila Abkhooie,2 Soheila Montazersaheb,3 Hossein Ghanbarian,4
    1. Molecular Medicine Research Center, Biomedicine institute, Tabriz University of Medical Sciences, Tabriz, Iran
    2. Department of Molecular Medicine, Faculty of Advanced Medical Sciences, Tabriz University of Medical Sciences, Tabriz, Iran
    3. Molecular Medicine Research Center, Biomedicine institute, Tabriz University of Medical Sciences, Tabriz, Iran
    4. Cellular and Molecular Biology Research Center, Shahid Beheshti University of Medical Sciences, Tehran, Iran


  • Introduction: Cdk9 is the catalytic core of the positive transcription elongation factor b (P-TEFb) and regulates transcriptional elongation factors by phosphorylation of RNA pol II. Apart from its role on myogenic gene expression, Cdk9 regulation of muscle-specific microRNAs in the early stage of cardiomyogenesis is poorly understood. Here we investigated potential regulatory functions of Cdk9 on myocardial microRNAs (MyomiRs) in myoblast C2C12 cells. Given the inhibitory role of microRNA-1 on Cdk9, we explored a potential negative regulatory feedback loop between microRNA-1 and the Cdk9 gene. We overexpressed Cdk9 in C2C12 myoblast cells and quantified myomiRs consist of: mir-1, miR-133 and miR-206 levels.
  • Methods: We overexpressed Cdk9 in mouse myoblast C2C12 cells to investigate its regulatory roles on myocardial microRNAs (MyomiRs). Upon Cdk9 overexpression, the expression level of important myomiRs consist of; miR-1, miR-133 and miR-206 were determined. Moreover, the expression profile of important muogenesis genes incluging MyoD, Mef2C, Srf and Myogenesis were examined in transfected and control cells by quantitative real‐time reverse transcription PCR (RT‐qPCR) and Western blot data analysis. In addition, Cell viability and apoptosis rate were determined via‐Annexin V FLUOS Staining Kit.
  • Results: Our resulted showed that, the significant induction of miR-1 and miR-206, while miR-133 was downregulated. Moreover, expression levels of MyoD and Srf, key regulators of myogenesis, increased in cells with overexpression of Cdk9. We further observed Cdk9-mediated apoptosis in C2C12 cells. MTT assays and annexin V/PI staining identified reduced proliferation, and a high level of apoptosis in Cdk9- transfected cells. Western blot analysis and quantitative RT-PCR determination of p53 and p21 showed a significant upregulation of these genes involved in apoptosis in Cdk9-overexpressing myocyte progenitor cells.
  • Conclusion: In agreement with Cdk9 regulatory functions on myomiRs, it has been proposed that Cdk9 promotes myoblast differentiation in the early stages of development. Increased expression of miR-1 and miR-206 and downregulation of miR-133 following over-expression of Cdk9 in transfected cells could promote differentiation of C2C12 cells. Furthermore, high expression levels of myogenic transcription factors, MyoD and Srf were observed in Cdk9-transfected cells. Whether the induction of miR-1 and miR-206 depends directly on Cdk9 or might be secondary to the induction of MyoD and Srf remains the subject of further studies. Based on our findings, we propose that Cdk9 controls myocyte progenitor cell growth, differentiation, and apoptosis by modulating myogenic transcription factors and myomiRs in a complex regulatory network.
  • Keywords: Myoblast cells, Cdk9, apoptosis, myomiRs and cardiac differentiation.