title: Role for the Ssu72 C-terminal domain phosphatase in RNA polymerase II transcription elongation.
abstract: The RNA polymerase II (RNAP II) transcription cycle is accompanied by changes in the phosphorylation status of the C-terminal domain (CTD), a reiterated heptapeptide sequence (Y(1)S(2)P(3)T(4)S(5)P(6)S(7)) present at the C terminus of the largest RNAP II subunit. One of the enzymes involved in this process is Ssu72, a CTD phosphatase with specificity for serine-5-P. Here we report that the ssu72-2-encoded Ssu72-R129A protein is catalytically impaired in vitro and that the ssu72-2 mutant accumulates the serine-5-P form of RNAP II in vivo. An in vitro transcription system derived from the ssu72-2 mutant exhibits impaired elongation efficiency. Mutations in RPB1 and RPB2, the genes encoding the two largest subunits of RNAP II, were identified as suppressors of ssu72-2. The rpb1-1001 suppressor encodes an R1281A replacement, whereas rpb2-1001 encodes an R983G replacement. This information led us to identify the previously defined rpb2-4 and rpb2-10 alleles, which encode catalytically slow forms of RNAP II, as additional suppressors of ssu72-2. Furthermore, deletion of SPT4, which encodes a subunit of the Spt4-Spt5 early elongation complex, also suppresses ssu72-2, whereas the spt5-242 allele is suppressed by rpb2-1001. These results define Ssu72 as a transcription elongation factor. We propose a model in which Ssu72 catalyzes serine-5-P dephosphorylation subsequent to addition of the 7-methylguanosine cap on pre-mRNA in a manner that facilitates the RNAP II transition into the elongation stage of the transcription cycle.
authors.affiliation: Department of Biochemistry, Robert Wood Johnson Medical School, 683 Hoes Lane West, Piscataway, NJ 08854, USA.
authors.forename: Mariela
authors.initials: M
authors.lastname: Reyes-Reyes
authors.forename: Michael
authors.initials: M
authors.lastname: Hampsey
mesh_terms: D000409:Alanine / Q000235:genetics; D000483:Alleles; D000595:Amino Acid Sequence; D001120:Arginine / Q000235:genetics; D002352:Carrier Proteins / Q000737:chemistry / Q000378:metabolism*; D002868:Chromosomal Proteins, Non-Histone / Q000378:metabolism; D004271:DNA, Fungal; D008957:Models, Genetic; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000378:metabolism; D010641:Phenotype; D010749:Phosphoprotein Phosphatases / Q000378:metabolism*; D011485:Protein Binding; D017433:Protein Structure, Secondary; D012319:RNA Polymerase II / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000166:cytology / Q000201:enzymology* / Q000254:growth & development; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry / Q000378:metabolism*; D017422:Sequence Analysis, DNA; D013489:Suppression, Genetic; D035381:TATA-Binding Protein Associated Factors / Q000378:metabolism; D035362:Transcription Factor TFIID; D014158:Transcription, Genetic; D035602:Transcriptional Elongation Factors / Q000378:metabolism; D039221:mRNA Cleavage and Polyadenylation Factors
chemical_list: D002352:Carrier Proteins; D002868:Chromosomal Proteins, Non-Histone; D004271:DNA, Fungal; D009687:Nuclear Proteins; C080991:SPT4 protein, S cerevisiae; C416934:SSU72 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; C087176:TAF2 protein, S cerevisiae; D035381:TATA-Binding Protein Associated Factors; D035362:Transcription Factor TFIID; D035602:Transcriptional Elongation Factors; D039221:mRNA Cleavage and Polyadenylation Factors; C069940:SPT5 transcriptional elongation factor; D001120:Arginine; D012319:RNA Polymerase II; C511736:RPB1 protein, S cerevisiae; D010749:Phosphoprotein Phosphatases; C116022:carboxy-terminal domain phosphatase; D000409:Alanine