title: How an mRNA capping enzyme reads distinct RNA polymerase II and Spt5 CTD phosphorylation codes.
abstract: Interactions between RNA guanylyltransferase (GTase) and the C-terminal domain (CTD) repeats of RNA polymerase II (Pol2) and elongation factor Spt5 are thought to orchestrate cotranscriptional capping of nascent mRNAs. The crystal structure of a fission yeast GTase•Pol2 CTD complex reveals a unique docking site on the nucleotidyl transferase domain for an 8-amino-acid Pol2 CTD segment, S5PPSYSPTS5P, bracketed by two Ser5-PO4 marks. Analysis of GTase mutations that disrupt the Pol2 CTD interface shows that at least one of the two Ser5-PO4-binding sites is required for cell viability and that each site is important for cell growth at 37°C. Fission yeast GTase binds the Spt5 CTD at a separate docking site in the OB-fold domain that captures the Trp4 residue of the Spt5 nonapeptide repeat T(1)PAW(4)NSGSK. A disruptive mutation in the Spt5 CTD-binding site of GTase is synthetically lethal with mutations in the Pol2 CTD-binding site, signifying that the Spt5 and Pol2 CTDs cooperate to recruit capping enzyme in vivo. CTD phosphorylation has opposite effects on the interaction of GTase with Pol2 (Ser5-PO4 is required for binding) versus Spt5 (Thr1-PO4 inhibits binding). We propose that the state of Thr1 phosphorylation comprises a binary "Spt5 CTD code" that is read by capping enzyme independent of and parallel to its response to the state of the Pol2 CTD.
authors.affiliation: Structural Biology Program, Sloan-Kettering Institute, New York, New York 10065, USA;
authors.forename: Selom K
authors.initials: SK
authors.lastname: Doamekpor
authors.affiliation: Microbiology and Immunology Department, Weill Cornell Medical College, New York, New York 10065, USA;
authors.forename: Ana M
authors.initials: AM
authors.lastname: Sanchez
authors.affiliation: Microbiology and Immunology Department, Weill Cornell Medical College, New York, New York 10065, USA;
authors.forename: Beate
authors.initials: B
authors.lastname: Schwer
authors.affiliation: Molecular Biology Program, Sloan-Kettering Institute, New York, New York 10065, USA;
authors.forename: Stewart
authors.initials: S
authors.lastname: Shuman
authors.affiliation: Structural Biology Program, Sloan-Kettering Institute, New York, New York 10065, USA; Howard Hughes Medical Institute, Structural Biology Program, Sloan-Kettering Institute, New York, New York 10065, USA.
authors.forename: Christopher D
authors.initials: CD
authors.lastname: Lima
mesh_terms: D005815:Genetic Code; D008958:Models, Molecular; D009713:Nucleotidyltransferases / Q000737:chemistry / Q000378:metabolism*; D010766:Phosphorylation; D020836:Protein Structure, Quaternary; D017434:Protein Structure, Tertiary; D012319:RNA Polymerase II / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012568:Schizosaccharomyces / Q000201:enzymology* / Q000235:genetics / Q000378:metabolism; D029702:Schizosaccharomyces pombe Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D013912:Threonine / Q000378:metabolism; D035602:Transcriptional Elongation Factors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D000098556:mRNA Guanylyltransferases
chemical_list: D029702:Schizosaccharomyces pombe Proteins; C549622:Spt5 protein, S pombe; D035602:Transcriptional Elongation Factors; D013912:Threonine; D009713:Nucleotidyltransferases; D012319:RNA Polymerase II; D000098556:mRNA Guanylyltransferases
keywords: RNA polymerase II; Spt5; mRNA capping; regulation; transcription