RESUMO
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.
Assuntos
Proteínas de Transporte/genética , Proteínas Fúngicas/metabolismo , RNA Polimerase II/metabolismo , Proteínas de Saccharomyces cerevisiae , Saccharomyces cerevisiae/metabolismo , Sequência de Aminoácidos , Proteínas de Transporte/fisiologia , Sequência Conservada , Proteínas Fúngicas/genética , Dados de Sequência Molecular , Mutação , Fosfoproteínas Fosfatases , Filogenia , Subunidades Proteicas , RNA Polimerase II/genética , Saccharomyces cerevisiae/genética , Supressão Genética , Fator de Transcrição TFIIB , Fatores de Transcrição/genética , Transcrição Gênica , Fatores de Poliadenilação e Clivagem de mRNARESUMO
Multiphoton resonance ionization has been combined with energetic ion bombardment to examine dopant concentrations ofindium on the surface of silicon. The results yield a linear relation between the indium concentration and the known bulk values and a detection limit of 9 parts per trillion, at a mass resolution exceeding 160. This measurement, which surpasses the limits of any previous surface analysis by a factor of 100, has been made possible with an experimental configuration that optimizes sampling and detection efficiency while reducing background noise to virtually zero. During the analysis, submonolayer quantities of the surface are removed, so that as few as 180 surface atoms may be counted.
