Tyr1 phosphorylation promotes phosphorylation of Ser2 on the C-terminal domain of eukaryotic RNA polymerase II by P-TEFb

eLife

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[1] Department of Chemical Engineering, University of Texas at Austin, Austin, United States;Department of Chemistry, University of Texas at Austin, Austin, United States;Department of Molecular Biosciences, University of Texas at Austin, Austin, United States;Department of Molecular Biosciences, University of Texas at Austin, Austin, United States;Institute for Cellular and Molecular Biology, University of Texas at Austin, Austin, United States;Department of Molecular Medicine, Institute of Biotechnology, University of Texas Health Science Center at San Antonio, San Antonio, United States;

关键词: post-translational modification;    P-TEFb;    transcription;    promoter-proximal pausing;    phosphorylation;    ultraviolet photodissociation mass spectrometry;    Human;   

DOI  :  10.7554/eLife.48725

来源: publisher

【 摘 要 】

10.7554/eLife.48725.001The Positive Transcription Elongation Factor b (P-TEFb) phosphorylates Ser2 residues of the C-terminal domain (CTD) of the largest subunit (RPB1) of RNA polymerase II and is essential for the transition from transcription initiation to elongation in vivo. Surprisingly, P-TEFb exhibits Ser5 phosphorylation activity in vitro. The mechanism garnering Ser2 specificity to P-TEFb remains elusive and hinders understanding of the transition from transcription initiation to elongation. Through in vitro reconstruction of CTD phosphorylation, mass spectrometry analysis, and chromatin immunoprecipitation sequencing (ChIP-seq) analysis, we uncover a mechanism by which Tyr1 phosphorylation directs the kinase activity of P-TEFb and alters its specificity from Ser5 to Ser2. The loss of Tyr1 phosphorylation causes an accumulation of RNA polymerase II in the promoter region as detected by ChIP-seq. We demonstrate the ability of Tyr1 phosphorylation to generate a heterogeneous CTD modification landscape that expands the CTD’s coding potential. These findings provide direct experimental evidence for a combinatorial CTD phosphorylation code wherein previously installed modifications direct the identity and abundance of subsequent coding events by influencing the behavior of downstream enzymes.

【 授权许可】

CC BY