Functional interaction between Ssu72 and the Rpb2 subunit of RNA polymerase II in Saccharomyces cerevisiae

Mol Cell Biol. 2000 Nov;20(22):8343-51. doi: 10.1128/MCB.20.22.8343-8351.2000.

Abstract

SSU72 is an essential gene encoding a phylogenetically conserved protein of unknown function that interacts with the general transcription factor TFIIB. A recessive ssu72-1 allele was identified as a synthetic enhancer of a TFIIB (sua7-1) defect, resulting in a heat-sensitive (Ts(-)) phenotype and a dramatic downstream shift in transcription start site selection. Here we describe a new allele, ssu72-2, that confers a Ts(-) phenotype in a SUA7 wild-type background. In an effort to further define Ssu72, we isolated suppressors of the ssu72-2 mutation. One suppressor is allelic to RPB2, the gene encoding the second-largest subunit of RNA polymerase II (RNAP II). Sequence analysis of the rpb2-100 suppressor defined a cysteine replacement of the phylogenetically invariant arginine residue at position 512 (R512C), located within homology block D of Rpb2. The ssu72-2 and rpb2-100 mutations adversely affected noninduced gene expression, with no apparent effects on activated transcription in vivo. Although isolated as a suppressor of the ssu72-2 Ts(-) defect, rpb2-100 enhanced the transcriptional defects associated with ssu72-2. The Ssu72 protein interacts directly with purified RNAP II in a coimmunoprecipitation assay, suggesting that the genetic interactions between ssu72-2 and rpb2-100 are a consequence of physical interactions. These results define Ssu72 as a highly conserved factor that physically and functionally interacts with the RNAP II core machinery during transcription initiation.

Publication types

  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Amino Acid Sequence
  • Carrier Proteins / genetics*
  • Carrier Proteins / physiology
  • Conserved Sequence
  • Fungal Proteins / genetics
  • Fungal Proteins / metabolism*
  • Molecular Sequence Data
  • Mutation
  • Phosphoprotein Phosphatases
  • Phylogeny
  • Protein Subunits
  • RNA Polymerase II / genetics
  • RNA Polymerase II / metabolism*
  • Saccharomyces cerevisiae / genetics
  • Saccharomyces cerevisiae / metabolism*
  • Saccharomyces cerevisiae Proteins*
  • Suppression, Genetic
  • Transcription Factor TFIIB
  • Transcription Factors / genetics
  • Transcription, Genetic
  • mRNA Cleavage and Polyadenylation Factors

Substances

  • Carrier Proteins
  • Fungal Proteins
  • Protein Subunits
  • SSU72 protein, S cerevisiae
  • Saccharomyces cerevisiae Proteins
  • Transcription Factor TFIIB
  • Transcription Factors
  • mRNA Cleavage and Polyadenylation Factors
  • RNA Polymerase II
  • Phosphoprotein Phosphatases