title: Direct Analysis of Phosphorylation Sites on the Rpb1 C-Terminal Domain of RNA Polymerase II.
abstract: Dynamic interactions between RNA polymerase II and various mRNA-processing and chromatin-modifying enzymes are mediated by the changing phosphorylation pattern on the C-terminal domain (CTD) of polymerase subunit Rpb1 during different stages of transcription. Phosphorylations within the repetitive heptamer sequence (YSPTSPS) of CTD have primarily been defined using antibodies, but these do not distinguish different repeats or allow comparative quantitation. Using a CTD modified for mass spectrometry (msCTD), we show that Ser5-P and Ser2-P occur throughout the length of CTD and are far more abundant than other phosphorylation sites. msCTD extracted from cells mutated in several CTD kinases or phosphatases showed the expected changes in phosphorylation. Furthermore, msCTD associated with capping enzyme was enriched for Ser5-P while that bound to the transcription termination factor Rtt103 had higher levels of Ser2-P. These results suggest a relatively sparse and simple "CTD code."
authors.affiliation: Department of Biochemical Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, MA 02115, USA; Department of Cancer Biology and Blais Proteomics Center, Dana Farber Cancer Institute, Boston, MA 02115, USA.
authors.forename: Hyunsuk
authors.initials: H
authors.lastname: Suh
authors.affiliation: Department of Biochemical Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, MA 02115, USA; Department of Cancer Biology and Blais Proteomics Center, Dana Farber Cancer Institute, Boston, MA 02115, USA.
authors.forename: Scott B
authors.initials: SB
authors.lastname: Ficarro
authors.affiliation: Department of Biochemical Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, MA 02115, USA; Department of Cancer Biology and Blais Proteomics Center, Dana Farber Cancer Institute, Boston, MA 02115, USA.
authors.forename: Un-Beom
authors.initials: UB
authors.lastname: Kang
authors.affiliation: Department of Biochemical Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, MA 02115, USA.
authors.forename: Yujin
authors.initials: Y
authors.lastname: Chun
authors.affiliation: Department of Biochemical Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, MA 02115, USA; Department of Cancer Biology and Blais Proteomics Center, Dana Farber Cancer Institute, Boston, MA 02115, USA.
authors.forename: Jarrod A
authors.initials: JA
authors.lastname: Marto
authors.affiliation: Department of Biochemical Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, MA 02115, USA. Electronic address: steveb@hms.harvard.edu.
authors.forename: Stephen
authors.initials: S
authors.lastname: Buratowski
mesh_terms: D000595:Amino Acid Sequence; D049109:Cell Proliferation; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D009154:Mutation; D010766:Phosphorylation; D017434:Protein Structure, Tertiary; D012319:RNA Polymerase II / Q000737:chemistry* / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000166:cytology / Q000201:enzymology* / Q000235:genetics; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry* / Q000378:metabolism*; D014158:Transcription, Genetic
chemical_list: D029701:Saccharomyces cerevisiae Proteins; D012319:RNA Polymerase II; C511736:RPB1 protein, S cerevisiae