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Characterization of a staurosporine- and temperature-sensitive mutant, stt1, of Saccharomyces cerevisiae: STT1 is allelic to PKC1

Molecular and General Genetics MGG volume 231, pages 337–344 (1992)Cite this article

Summary

Staurosporine is an antibiotic that specifically inhibits protein kinase C. Fourteen staurosporine- and temperature-sensitive (stt) mutants of Saccharomyces cerevisiae were isolated and characterized. These mutants were divided into ten complementation groups, and characterized for their cross-sensitivity to K-252a, neomycin, or CaCl2, The STT1 gene was cloned and sequenced. The nucleotide sequence of the STT1 gene revealed that STT1 is the same gene as PKC1. The STT1 gene conferred resistance to staurosporine on wild-type cells, when present on a high copy number plasmid. STT1/stt1::HIS3 diploid cells were more sensitive to staurosporine than STT1/STT1 diploid cells. Analysis of temperature-sensitive stt1 mutants showed that the STT1 gene product functioned in S or G2/M phase. These results suggest that a protein kinase (the STT1 gene product) is one of the essential targets of staurosporine in yeast cells.

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References

  • Abe K, Yoshida M, Usui T, Horinouchi S, Beppu T (1991) High synchronous culture of fibroblasts from G2 block caused by staurosporine, a potent inhibitor of protein kinase. Exp Cell Res 192:122–127

    Google Scholar 

  • Adams AEM, Johnson DI, Longnecker RM, Sloat BF, Pringle JR (1990) CDC42 and CDC43, two additional genes involved in budding and the establishment of cell polarity in the yeast Saccharomyces cerevisiae. J Cell Biol 111:131–142

    Google Scholar 

  • Becker GW, Lester RL (1977) Changes in phospholipids of Saccharomyces cerevisiae associated with inositol-less death. J Biol Chem 252:8684–8690

    Google Scholar 

  • Belunis DJ, Bae-Lee M, Kelley MJ, Carman GM (1988) Purification and characterization of phosphatidylinositol kinase from Saccharomyces cerevisiae. J Biol Chem 263:18897–18903

    Google Scholar 

  • Berridge MJ (1987) Inositol trisphosphate and diacylglycerol: two interacting second messengers. Annu Rev Biochem 56:159–193

    Google Scholar 

  • Bullock WO, Fernandez JM, Short JM (1987) A high efficiency plasmid transforming recA Escherichia coli strain with beta galactosidase selection. BioTechniques 5:376–379

    Google Scholar 

  • Castagna M, Takai Y, Kaibuchi K, Sano K, Kikkawa U, Nishizuka Y (1982) Direct activation of calcium-activated, phospholipid-dependent protein kinase by tumor-promoting phorbol esters. J Biol Chem 257:7847–7851

    Google Scholar 

  • Hawthorne DC, Friis J (1964) Osmotic-remedial mutants. A new classification for nutritional mutants in yeast. Genetics 50:829–839

    Google Scholar 

  • Henry SA (1982) Membrane lipids of yeast: biochemical and genetic studies. In Strathern JN, Jones EW, Broach JR (eds) The molecular biology of the yeast Saccharomyces: metabolism and gene expression. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY, pp 101–158

    Google Scholar 

  • Hill JE, Myers AM, Koerner TJ, Tzagoloff A (1986) Yeast/E. coli shuttle vectors with multiple unique restriction sites. Yeast 2:163–167

    Google Scholar 

  • Hutter K-J, Eipel HE (1979) DNA determination of yeast by flow cytometry. J Gen Microbiol 113:369–375

    Google Scholar 

  • Iida H, Sakaguchi S, Yagawa Y, Anraku Y (1990) Cell cycle control by Ca2+ in Saccharomyces cerevisiae. J Biol Chem 265:21216–21222

    Google Scholar 

  • Ito H, Fukuda Y, Murata K, Kimura A (1983) Transformation of intact yeast cells treated with alkali cations. J Bacteriol 153:163–168

    Google Scholar 

  • Jones JS, Prakash L (1990) Yeast Saccharomyces cerevisiae selectable markers in pUC18 polylinkers. Yeast 6:363–366

    Google Scholar 

  • Kase H, Iwahashi K, Matsuda Y (1986) K-252a, a potent inhibitor of protein kinase C from microbial origin. J Antibiot (Tokyo) 39:1059–1065

    Google Scholar 

  • Kohno T, Roth J (1979) Electrolytic effects on the activity of mutant enzymes in vivo and in vitro. Biochemistry 18:1386–1392

    Google Scholar 

  • Lester RL, Steiner MR (1968) The occurrence of diphosphoinositide and triphosphoinositide in Saccharomyces cerevisiae. J Biol Chem 243:4889–4893

    Google Scholar 

  • Levin DE, Fields FO, Kunisawa R, Bishop JM, Thorner J (1990) A candidate protein kinase C gene (PKC1) is required for the S. cerevisiae cell cycle. Cell 62:213–224

    Google Scholar 

  • Lovett ST, Mortimer RK (1987) Characterization of null mutants of the RAD55 gene of Saccharomyces cerevisiae: effects of temperature, osmotic strength and mating type. Genetics 116:547–553

    Google Scholar 

  • Maniatis T, Fritsch EF, Sambrook J (1982) Molecular cloning: a laboratory manual. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY

    Google Scholar 

  • Matsumoto K, Uno I, Ishikawa T (1986) Fluphenazine-resistant Saccharomyces cerevisiae mutants defective in the cell division cycle. J Bacteriol 168:1352–1357

    Google Scholar 

  • Nakano H, Kobayashi E, Takahashi I, Tamaoki T, Kuzuu Y, Iba H (1987) Staurosporine inhibits tyrosine-specific protein kinase activity of Rous sarcoma virus transforming protein p60. J Antibiot (Tokyo) 40:706–708

    Google Scholar 

  • Nasmyth KA, Reed SI (1980) Isolation of genes by complementation in yeast: molecular cloning of a cell-cycle gene. Proc Natl Acad Sci USA 77:2119–2123

    Google Scholar 

  • Nikawa J, Kodaki T, Yamashita S (1987) Primary structure and disruption of the phosphatidylinositol synthase gene of Saccharomyces cerevisiae. J Biol Chem 262:4876–4881

    Google Scholar 

  • Nishizuka Y (1984a) Turnover of inositol phospholipids and signal transduction. Science 225:1365–1370

    Google Scholar 

  • Nishizuka Y (1984b) The role of protein kinase C in cell surface signal transduction and tumor promoter. Nature 308:693–698

    Google Scholar 

  • Ogita K, Miyamoto S, Koide H, Iwai T, Oka M, Ando K, Kishimoto A, Ikeda K, Fukami Y, Nishizuka Y (1990) Protein kinase C in Saccharomyces cerevisiae: comparison with the mammalian enzyme. Proc Natl Acad Sci USA 87:5011–5015

    Google Scholar 

  • Ohya Y, Ohsumi Y, Anraku Y (1984) Genetic study of the role of calcium ions in the cell division cycle of Saccharomyces cerevisiae: a calcium-dependent mutant and its trifluoperazine-dependent pseudorevertants. Mol Gen Genet 193:389–394

    Google Scholar 

  • Omura S, Iwai Y, Hirano A, Nakagawa A, Awaya J, Tsuchiya H, Takahashi Y, Masuma R (1977) A new alkaloid AM-2282 of Streptomyces origin taxonomy, fermentation, isolation and preliminary characterization. J Antibiot (Tokyo) 30:275–282

    Google Scholar 

  • Sanger F, Nicklen S, Coulson AR (1977) DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci USA 83:5463–5467

    Google Scholar 

  • Schmitt HD, Puzicha M, Gallwitz D (1988) Study of a temperature-sensitive mutant of the ras-related YPT1 gene product in yeast suggests a role in the regulation of intracellular calcium. Cell 53:635–647

    Google Scholar 

  • Sherman F, Fink GR, Hicks JB (1986) Methods in yeast genetics. Cold Spring Harbor Laboratory, Cold Spring Harbor, New York

    Google Scholar 

  • Shimma Y, Uno I (1990) Isolation and characterization of neomycin-sensitive mutants in Saccharomyces cerevisiae. J Gen Microbiol 136:1753–1761

    Google Scholar 

  • Sikorski RS, Hieter P (1989) A system of shuttle vectors and yeast host strains designed for efficient manipulation of DNA in Saccharomyces cerevisiae. Genetics 122:19–27

    Google Scholar 

  • Streb H, Irvine RF, Berridge MJ, Schulz I (1983) Release of Ca2+ from a nonmitochondrial intracellular store in pancreatic acinar cells by inositol-1, 4, 5-trisphosphate. Nature 306:67–69

    Google Scholar 

  • Tamaoki T, Nomoto H, Takahashi I, Kato Y, Morimoto M, Tomita F (1986) Staurosporine, a potent inhibitor of phospholipid Ca+− dependent protein kinase. Biochem Biophys Res Commun 135:397–402

    Google Scholar 

  • Toda T, Shimanuki M, Yanagida M (1991) Fission yeast gene that confer resistance to stuarosporine encode an AP-1-like transcription factor and a protein kinase related to the mammalian ERK1/MAP2 and budding yeast FUS3 and KSS1 kinases. Genes Dev 5:60–73

    Google Scholar 

  • Winston F, Chumley F, Fink GR (1983) Genetic applications of yeast transformation with linear and gapped plasmids. Methods Enzymol 101:211–228

    Google Scholar 

  • Yasuzawa T, Iida T, Yoshida M, Hirayama N, Takahashi M, Shirahata K, Sano H (1986) The structure of novel protein kinase C inhibitor K-252a, b, c and d. J Antiobiot (Tokyo) 39:1072–1078

    Google Scholar 

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Author notes

  1. Satoshi Yoshida & Eri Ikeda

    Present address: Department of Botany, Faculty of Science, University of Tokyo, Bunkyo-ku, 113, Tokyo, Japan

Affiliations

  1. Institute of Applied Microbiology, University of Tokyo, Bunkyo-ku, 113, Tokyo, Japan

    Satoshi Yoshida, Eri Ikeda, Isao Uno & Hiroshi Mitsuzawa

Authors
  1. Satoshi Yoshida
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  2. Eri Ikeda
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  3. Isao Uno
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  4. Hiroshi Mitsuzawa
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Communicated by C. Hollenberg

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Yoshida, S., Ikeda, E., Uno, I. et al. Characterization of a staurosporine- and temperature-sensitive mutant, stt1, of Saccharomyces cerevisiae: STT1 is allelic to PKC1 . Molec. Gen. Genet. 231, 337–344 (1992). https://doi.org/10.1007/BF00292700

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  • DOI: https://doi.org/10.1007/BF00292700

Key words

  • Saccharomyces cerevisiae
  • Staurosporine
  • Protein kinase C
  • Cell cycle