PHOSPHORYLATION OF NUCLEAR MYOD IS REQUIRED FOR ITS RAPID DEGRADATION

MyoD is a basic helix-loop-helix transcription factor involved in the activation of genes encoding skeletal muscle-specific proteins. Indepe ndent of its ability to transactivate muscle-specific genes, MyoD can also act as a cell cycle inhibitor. MyoD activity is regulated by tran scriptional and posttranscriptional mechanisms. While MyoD can be foun d phosphorylated, the functional significance of this posttranslation modification has not been established. MyoD contains several consensus cyclin-dependent kinase (CDK) phosphorylation sites. In these studies , we examined whether a link could be established between MyoD activit y and phosphorylation at putative CDK sites. Site-directed mutagenesis of potential CDK phosphorylation sites in MyoD revealed that S200 is required for MyoD hyperphosphorylation as well as the normally short h alf-life of the MyoD protein. Additionally, we determined that turnove r of the MyoD protein requires the proteasome and Cdc34 ubiquitin-conj ugating enzyme activity. Results of these studies demonstrate that hyp erphosphorylated MyoD is targeted for rapid degradation by the ubiquit in pathway. The targeted degradation of MyoD following CDK phosphoryla tion identifies a mechanism through which MyoD activity can be regulat ed coordinately with the fell cycle machinery (CDK2 and CDK4) and/or c oordinately with the cellular transcriptional machinery (CDK7, CDK8, a nd CDK9).