Role of CDK9 in skeletal muscle

Cdk9 is a member of cyclin-dependent kinases and it is expressed in human and murine tissue with high levels in terminally differentiated cells. The elevated levels of Cdk9 and its regulatory subunits in terminally differentiated cells, together with the fact that it is implicated in the regulation of transcriptional elongation via phosphorylation of CTD of RNA polII, distinguishes Cdk9 from the other cdks. Cyclin partners of CDK9 are cyclin T1, T2a, T2b and K, with a formation of a complex, called p-TEF-B. In Hela cells about 80% of cellular P-TEFb is composed of cyclin T1 and the cyclin –dependent kinase 9 (CDK9), and about 20% of cellular CDK9 is complexed to other cyclins, such as cyclin T2a, T2b and with cyclin K. It was demonstrated that CdK9 complexed with CyT2a plays a role in the activation of myogenic program and Cdk9-CyT2a activity is not down-regulated in myotube formation, but its activation contributes to the transcriptional activity MyoD-mediated during myogenic program. The formation of a multimeric complex, containing cdk9/cyclin T2a and MyoD is present in muscle cells during the activation of the differentiation program and the N-terminal region of Cdk9 directly interacts with the b-HLH region of MyoD, allowing the formation of a complex also containing cyclin T2a. Recently a 55 kDa protein called CDK9-55 has been identified. This isoform presents 117 additional amminoacids residues at the N-terminal portion of Cdk9-42 and it conserves all molecular features of 42 isoform, indeed it associates with CycT phosphorylates CTD of RNA polII. Cdk9-55 is significantly upregulated in cells induced to differentiate, either in C2C12 cells or in satellite isolated cells, and it was demonstrated that there is a clear induction of cdk9-55 expression in injured skeletal muscles. Analysis in vivo on limbs at different times of development demostrated a different correlation between two CDK9 isoforms: CDK9-55 is involved during foetal myogenesis, whereas the CDK9-42 is expressed during embrional myogenesis. Study in vitro demonstrated that CDK9-55 is the isoform more expressed during differentiation both embrional and fetal myoblasts. A new antibody, p-CDK9, permitted us to define CDK9-55 as the active isoform during the differentiation. CDK9-55 is also the isoform more expressed during the re-innervation process. The analysis of different cyclins that complex with different members of kinase proteins showed that in vivo only cyclin K and D3 are involved during foetal myogenesis, suggesting their interaction with CDK9-55, whereas Cyclin T1, T2, L and H are implicated during primary myogenesis, in concomitance with CDK9-42 expression. Study in vitro demonstrated that only cyclin T2 is involved during the differentiation process for embrional and fetal myoblasts. Between inhibitory post transcriptional regulator mechanism of CDK9 there is the interaction with 7SKsnRNA and HEXIM1, our studies, on C2C12 cells, by RIP assay demonstrated that CDK9 isn’t associated with 7SKsnRNA/Hexim1 during differentiative phase, supporting that in differentiation CDK9 is active.