title: Serine-7 but not serine-5 phosphorylation primes RNA polymerase II CTD for P-TEFb recognition.
abstract: Phosphorylation of RNA polymerase II carboxy-terminal domain (CTD) in hepta-repeats YSPTSPS regulates eukaryotic transcription. Whereas Ser5 is phosphorylated in the initiation phase, Ser2 phosphorylation marks the elongation state. Here we show that the positive transcription elongation factor P-TEFb is a Ser5 CTD kinase that is unable to create Ser2/Ser5 double phosphorylations, while it exhibits fourfold higher activity on a CTD substrate pre-phosphorylated at Ser7 compared with the consensus hepta-repeat or the YSPTSPK variant. Mass spectrometry reveals an equal number of phosphorylations to the number of hepta-repeats provided, yet the mechanism of phosphorylation is distributive despite the repetitive nature of the substrate. Inhibition of P-TEFb activity is mediated by two regions in Hexim1 that act synergistically on Cdk9 and Cyclin T1. HIV-1 Tat/TAR abrogates Hexim1 inhibition to stimulate transcription of viral genes but does not change the substrate specificity. Together, these results provide insight into the multifaceted pattern of CTD phosphorylation.
authors.affiliation: Max-Planck-Institut für molekulare Physiologie, Abt. Physikalische Biochemie, Otto-Hahn-Straße 11, 44227 Dortmund, Germany.
authors.forename: Nadine
authors.initials: N
authors.lastname: Czudnochowski
authors.forename: Christian A
authors.initials: CA
authors.lastname: Bösken
authors.forename: Matthias
authors.initials: M
authors.lastname: Geyer
mesh_terms: D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D006801:Humans; D008969:Molecular Sequence Data; D010766:Phosphorylation; D042925:Positive Transcriptional Elongation Factor B / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D012319:RNA Polymerase II / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D016601:RNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D012694:Serine / Q000235:genetics / Q000378:metabolism*; D014157:Transcription Factors
chemical_list: C476123:HEXIM1 protein, human; D016601:RNA-Binding Proteins; D014157:Transcription Factors; D012694:Serine; D042925:Positive Transcriptional Elongation Factor B; D012319:RNA Polymerase II