分子细胞生物学前沿-博士-2010教程方案.ppt

分子细胞生物学专题----转录延伸因子P-TEFb及其活性调控 生命科学学院 陈瑞川 Main Points Transcription process of eukaryotic gene. P-TEFb: functions and its Activity regulation Steps of class II gene transcription RNA polymerase II (Pol II) P-TEFb: composition and property Discovered in 1995. (positive transcription elongation factor b) A heterodimer of Cdk9 and cyclin T, including T1, T2a, T2b and Cyclin K. But 80% P-TEFb is CDK9/Cyclin T1. kinase, phosphorylates CTD of Pol II, DSIF and NELF. How does cell signaling control the disruption of the inactive complex? Control P-TEFb activity by calcium signaling pathway UV and HMBA can trigger Ca+2 influx Calcium signaling pathways Calcium signaling pathways Hiding and seeking …. ? isoform of PP1 catalytic subunit involves in mediating the disruption of 7SK snRNP In vivo:PP2B and PP1 work cooperatively to disrupt the inactive complex In vitro: PP2B and PP1 directly disrupt inactive complex PP2B:let me work first…. How do PP2B and PP1 cooperatively disrupt inactive P-TFb complex? In vitro….Only PP1 can dephosphorylate pT186 of CDK9 In vitro….PP2B and PP1 work cooperatively to dephosphorylate pT186 of CDK9 How is liberated P-TFFb Recruited to chromatin Modified Nuclear Fractionation Almost all of Brd4 associate with chromatin at basal state Stimuation induces Brd4 release from chromatin The release of Brd4 from chromatin is essential from P-TEFb recruitment The release of Brd4 from chromatin is HDAC-dependent The release of Brd4 from chromatin is essential from elongation F1C2 cell Signal pathway inhibitors CycT1 Cdk9 HEXIM1 7SK snRNP Inactive P-TEFb P P P P P P P P P P P P P P P P Stress treatment CycT1 Cdk9 Active P-TEFb P P P P P P P HEXIM1 + A calcium-dependent signaling pathway controls P-TEFb transcriptional activity (in writing) Inhibitor: UV: HEXIM1 CDK9-f 1 2 3 4 5 Ca++ Influx CycT1 Cdk9 HEXIM1 7SK snRNP Inactive P-TEFb P P P P P P P P P P P P P P P P CycT1 Cdk9 Active P-TEFb P P P P P P P HEXIM1 + Cell memb