P-TEFb is a crucial co-factor for Myc transactivation.
Myc forms an heterodimer with Max and operates as a transcription factor upon binding to specific DNA sites in cellular chromatin. In addition to recruit histone acetylation activity, Myc binds to the positive transcription elongation factor b (P-TEFb) which consists of the cyclin-dependent kinase CKD9 and its regulatory subunit cyclin T. P-TEFb phosphorylates the carboxyl-terminal-domain (CTD) of the larger subunit of RNA polymerase II as well as negative elongation factors allowing efficient transcription elongation. Here, we report that Myc binds, as heterodimer with Max, exclusively the core active P-TEFb complex, and it recruits P-TEFb at Myc targets in vivo. Pharmacological inhibition of P-TEFb by 5.6-di-chloro-1-b-D-ribofuranosyl-bensimidazole (DRB) specifically inhibits expression of Myc-responsive CAD and NUC genes, and impairs the Myc-induced S-phase and apoptosis of quiescent cells grown in low serum. Chromatin immunoprecipitation assays (ChIP) demonstrated co-occupancy of Myc and P-TEFb to CAD and NUC E-boxes, and DRB treatment diminished the density of Pol II phosphorylated on Ser-2 of its CTD. These results indicate that P-TEFb is recruited in vivo to Myc-target promoters and CDK9 activity is an important step for Myc-dependent stimulation of responsive genes.
Gargano
Barbara
B
Department of Structural and Functional Biology, University of Naples Federico II, Naples, Italy.
Amente
Stefano
S
Majello
Barbara
B
Lania
Luigi
L
D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D002453:Cell Cycle / Q000187:drug effects; D049109:Cell Proliferation / Q000187:drug effects; D002478:Cells, Cultured; D047369:Chromatin Immunoprecipitation; D042863:Cyclin-Dependent Kinase 9 / Q000235:genetics / Q000378:metabolism*; D004004:Dichlororibofuranosylbenzimidazole / Q000494:pharmacology; D015870:Gene Expression / Q000187:drug effects; D010766:Phosphorylation / Q000187:drug effects; D042925:Positive Transcriptional Elongation Factor B / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D011401:Promoter Regions, Genetic / Q000235:genetics; D011485:Protein Binding / Q000187:drug effects; D021122:Protein Subunits / Q000235:genetics / Q000378:metabolism; D016271:Proto-Oncogene Proteins c-myc / Q000235:genetics / Q000378:metabolism*; D012319:RNA Polymerase II / Q000235:genetics / Q000378:metabolism; D051381:Rats; D020133:Reverse Transcriptase Polymerase Chain Reaction
D021122:Protein Subunits; D016271:Proto-Oncogene Proteins c-myc; D004004:Dichlororibofuranosylbenzimidazole; D042925:Positive Transcriptional Elongation Factor B; D042863:Cyclin-Dependent Kinase 9; D012319:RNA Polymerase II