title: Evidence that P-TEFb alleviates the negative effect of DSIF on RNA polymerase II-dependent transcription in vitro.
abstract: Recently, a positive and a negative elongation factor, implicated in 5,6-dichloro-1-beta-D-ribofuranosylbenzimidazole (DRB) inhibition of transcription elongation, has been identified. P-TEFb is a positive transcription elongation factor and the DRB-sensitive kinase that phosphorylates the C-terminal domain (CTD) of the largest subunit of RNA polymerase II (Pol II). PITALRE, a member of the Cdc2 family of protein kinases, is the catalytic subunit of P-TEFb. DSIF is a human homolog of the yeast Spt4-Spt5 complex and renders elongation of transcription sensitive to DRB. DRB sensitivity-inducing factor (DSIF) binds to RNA Pol II and may directly regulate elongation. Here we show a functional interaction between P-TEFb and DSIF. The reduction of P-TEFb activity induced by either DRB, antibody against PITALRE, or immunodepletion resulted in a negative effect of DSIF on transcription. DSIF acts at an early phase of elongation, and the prior action of P-TEFb makes transcription resistant to DSIF. The state of phosphorylation of CTD determines the DSIF-RNA Pol II interaction, and may provide a direct link between P-TEFb and DSIF. Taken together, this study reveals a molecular basis for DRB action and suggests that P-TEFb stimulates elongation by alleviating the negative action of DSIF.
authors.affiliation: Faculty of Bioscience and Biotechnology, Tokyo Institute of Technology, 4259 Nagatsuta-cho, Midori-ku, Yokohama 226-8501, Japan.
authors.forename: T
authors.initials: T
authors.lastname: Wada
authors.forename: T
authors.initials: T
authors.lastname: Takagi
authors.forename: Y
authors.initials: Y
authors.lastname: Yamaguchi
authors.forename: D
authors.initials: D
authors.lastname: Watanabe
authors.forename: H
authors.initials: H
authors.lastname: Handa
mesh_terms: D042863:Cyclin-Dependent Kinase 9; D018844:Cyclin-Dependent Kinases / Q000378:metabolism; D004004:Dichlororibofuranosylbenzimidazole / Q000494:pharmacology; D005786:Gene Expression Regulation; D006801:Humans; D008957:Models, Genetic; D009687:Nuclear Proteins / Q000378:metabolism*; D010766:Phosphorylation; D042925:Positive Transcriptional Elongation Factor B; D011494:Protein Kinases; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D012319:RNA Polymerase II / Q000378:metabolism*; D012333:RNA, Messenger / Q000096:biosynthesis; D012097:Repressor Proteins; D014157:Transcription Factors; D014158:Transcription, Genetic; D035602:Transcriptional Elongation Factors
chemical_list: D009687:Nuclear Proteins; D012333:RNA, Messenger; D012097:Repressor Proteins; C481011:SUPT4H1 protein, human; C112224:SUPT5H protein, human; D014157:Transcription Factors; D035602:Transcriptional Elongation Factors; D004004:Dichlororibofuranosylbenzimidazole; D011494:Protein Kinases; C060137:carboxy-terminal domain kinase; D042925:Positive Transcriptional Elongation Factor B; D017346:Protein Serine-Threonine Kinases; C467969:CDK9 protein, human; D042863:Cyclin-Dependent Kinase 9; D018844:Cyclin-Dependent Kinases; D012319:RNA Polymerase II