Multiple cellular processes affected by the absence of the Rpb4 subunit of RNA polymerase II contribute to the deficienc
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O R I GI N A L P A P E R
Y. Bourbonnais á N. Faucher á D. Pallotta á C. Larouche
Multiple cellular processes affected by the absence of the Rpb4 subunit of RNA polymerase II contribute to the de®ciency in the stress response of the yeast rpb4D mutant Received: 28 June 2000 / Accepted: 23 August 2000 / Published online: 11 October 2000 Ó Springer-Verlag 2000
Abstract We previously described the isolation of yeast mutants (sex mutants) that secrete reduced amounts of mature a-factor when it is synthesized as part of a fusion with prosomatostatin. In the present study we show that the sex3-1 mutant displays pleiotropic phenotypes. These include an abnormal morphology, an osmoremediable caeine sensitivity, reduced secretion of mature a-factor, a weakened cell wall and a marked de®ciency in halotolerance. Cloning of the SEX3 gene revealed that it is identical to the RPB4 gene. This gene encodes the fourth largest subunit of yeast RNA polymerase II, which has been postulated to play a major role in the response to stress. We show that transcriptional activation in response to either a cell wall stress or to growth in the presence of elevated salt concentrations is minimally aected by the loss of RPB4 function. However, whereas the levels of several mRNAs are similarly reduced (by about 30%) in rpb4 mutants grown in rich medium at moderate temperature, some transcripts, in particular ZDS1, are more abundant. An increase dosage of ZDS1, or of genes involved in cell wall assembly and in secretion (RHO1 and SRO77, respectively), partially suppresses the sensitivity of rpb4D cells to high temperature, heat shock and stationary phase. Collectively, our results indicate that the loss of Rpb4p perturbs several cellular functions that contribute to the inappropriate stress response of rpb4D
Communicated by D. Y. Thomas Y. Bourbonnais (&) á C. Larouche DeÂpartement de biochimie et de microbiologie, Centre de Recherche sur la Fonction, la Structure et l'IngeÂnierie des ProteÂines (CREFSIP), Pavillon Charles-EugeÁne Marchand, Universite Laval, QueÂbec, Canada G1K 7P4 E-mail: [email protected] N. Faucher á D. Pallotta DeÂpartement de biologie, Centre de Recherche sur la Fonction, la Structure et l'IngeÂnierie des ProteÂines (CREFSIP), Pavillon Charles-EugeÁne Marchand, Universite Laval, QueÂbec, Canada G1K 7P4
yeast. We therefore conclude that this RNA polymerase II subunit is not speci®cally involved in the stress response. Key words RNA polymerase II subunit á Saccharomyces cerevisiae á Transcription á Genetic suppressors á Cell wall integrity
Introduction RNA polymerase II is a complex enzyme whose subunits are highly conserved among eukaryotes. In the yeast S. cerevisiae, the core enzyme is composed of 12 subunits with molecular sizes varying from 10 kDa up to 220 kDa. By a combination of methods, including screening of yeast genomic DNA with cDNAs from other species, and the use of subunit-speci®c antibodies and oligonucleotide probes derived from amino acid sequences of individual subunits, the genes encod
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