RESUMO
SSC1 is an essential member of the yeast HSP70 multigene family (E. Craig, J. Kramer, and J. Kosic-Smithers, Proc. Natl. Acad. Sci. USA 84:4156-4160, 1987). Analysis of the SSC1 DNA sequence revealed that it could encode a 70,627-dalton protein that is more similar to DnaK, an Escherichia coli hsp70 protein, than other yeast hsp70s whose sequences have been determined. Ssc1p was found to have an amino-terminal extension of 28 amino acids, in comparison with either Ssa1p, another hsp70 yeast protein, or Dnak. This putative leader is rich in basic and hydroxyl amino acids, characteristic of many mitochondrial leader sequences. Ssc1p that was synthesized in vitro could be imported into mitochondria and was cleaved in the process. The imported protein comigrated with an abundant mitochondrial protein that reacted with hsp70-specific antibodies. We conclude that Ssc1p is a mitochondrial protein and that hsp70 proteins perform functions in many compartments of the cell.
Assuntos
Proteínas Fúngicas/genética , Genes Fúngicos , Proteínas de Choque Térmico/genética , Mitocôndrias/metabolismo , Família Multigênica , Saccharomyces cerevisiae/genética , Sequência de Aminoácidos , Sequência de Bases , Transporte Biológico , Western Blotting , Cromatografia em Gel , DNA Fúngico/genética , Escherichia coli/genética , Proteínas Fúngicas/metabolismo , Proteínas de Choque Térmico/metabolismo , Dados de Sequência Molecular , Plasmídeos , Sinais Direcionadores de Proteínas/genética , Sinais Direcionadores de Proteínas/metabolismoRESUMO
Yeast Hsp70 genes constitute a multigene family in which at least five of the nine members are heat inducible. Hsp70 RNA levels also vary dramatically during stationary arrest and sporulation. During growth to stationary phase, SSB1-SSB2 and SSC1 RNAs decreased in abundance as cell density increased. In contrast, SSA1-SSA2 RNA levels increased before the diauxic shift and then decreased as cells approach stationary phase. SSA3 RNA was detected only after the diauxic shift and accumulated to high levels as cells entered stationary phase. This accumulation was reversed by addition of glucose. Studies with cyr1 mutants indicated that SSA3 RNA accumulation is stimulated by decreasing intracellular cyclic AMP concentrations. When cells were incubated in sporulation medium, most Hsp70 RNAs, with the exception of SSA1-SSA2 RNA, decreased in abundance. This finding contrasted with the SSA1-SSA2 pattern observed during growth to stationary phase. SSA3 RNA was not detected during growth in acetate-based medium but accumulated after several hours. SSA3 RNA accumulation was higher in sporulating cells than in nonsporulating cells and was reversed by addition of glucose.
Assuntos
Proteínas de Choque Térmico/genética , Saccharomyces cerevisiae/fisiologia , Eletroforese em Gel Bidimensional , Regulação da Expressão Gênica , Temperatura Alta , Cinética , RNA Fúngico/metabolismo , RNA Mensageiro/metabolismo , Esporos FúngicosRESUMO
The genome of the yeast Saccharomyces cerevisiae contains a family of genes related to the HSP70 genes (encoding the 70-kDa heat shock protein) of other eukaryotes. Mutations in two of these yeast genes (SSC1 and SSD1), whose expression is increased a few fold after temperature upshift, were constructed in vitro and substituted into the yeast genome in place of the wild-type alleles. No phenotypic effects of the mutation in SSD1 were detected. However, a functional SSC1 gene is essential for vegetative growth. This result, in conjunction with experiments involving mutations in other members of this multigene family, indicates that at least three distinct functions are carried out by genes of the HSP70 family.
