POB3 is required for both transcription and replication in the yeast Saccharomyces cerevisiae.
Spt16 and Pob3 form stable heterodimers in Saccharomyces cerevisiae, and homologous proteins have also been purified as complexes from diverse eukaryotes. This conserved factor has been implicated in both transcription and replication and may affect both by altering the characteristics of chromatin. Here we describe the isolation and properties of a set of pob3 mutants and confirm that they have defects in both replication and transcription. Mutation of POB3 caused the Spt(-) phenotype, spt16 and pob3 alleles displayed severe synthetic defects, and elevated levels of Pob3 suppressed some spt16 phenotypes. These results are consistent with previous reports that Spt16 and Pob3 act in a complex that modulates transcription. Additional genetic interactions were observed between pob3 mutations and the genes encoding several DNA replication factors, including POL1, CTF4, DNA2, and CHL12. pob3 alleles caused sensitivity to the ribonucleotide reductase inhibitor hydroxyurea, indicating a defect in a process requiring rapid dNTP synthesis. Mutation of the S phase checkpoint gene MEC1 caused pob3 mutants to lose viability rapidly under restrictive conditions, revealing defects in a process monitored by Mec1. Direct examination of DNA contents by flow cytometry showed that S phase onset and progression were delayed when POB3 was mutated. We conclude that Pob3 is required for normal replication as well as for transcription.
Department of Biochemistry, University of Utah School of Medicine, Salt Lake City, Utah 84132, USA.
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Schlesinger
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D000483:Alleles; D002352:Carrier Proteins / Q000235:genetics* / Q000502:physiology*; D002453:Cell Cycle / Q000235:genetics; D018797:Cell Cycle Proteins / Q000235:genetics / Q000502:physiology; D002467:Cell Nucleus / Q000235:genetics; D002843:Chromatin / Q000378:metabolism; D004261:DNA Replication / Q000235:genetics*; D004591:Electrophoresis, Polyacrylamide Gel; D005434:Flow Cytometry; D006918:Hydroxyurea / Q000494:pharmacology; D016296:Mutagenesis; D009154:Mutation; D019384:Nucleic Acid Synthesis Inhibitors / Q000494:pharmacology; D016196:S Phase / Q000235:genetics; D012441:Saccharomyces cerevisiae / Q000235:genetics* / Q000502:physiology; D029701:Saccharomyces cerevisiae Proteins; D013696:Temperature; D013997:Time Factors; D014157:Transcription Factors / Q000378:metabolism; D014158:Transcription, Genetic / Q000235:genetics*; D035602:Transcriptional Elongation Factors
D002352:Carrier Proteins; D018797:Cell Cycle Proteins; D002843:Chromatin; D019384:Nucleic Acid Synthesis Inhibitors; C106768:POB3 protein, S cerevisiae; C071238:SPT16 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D014157:Transcription Factors; D035602:Transcriptional Elongation Factors; D006918:Hydroxyurea