Molecular and General Genetics MGG

, Volume 244, Issue 5, pp 519–529 | Cite as

The vacuolar compartment is required for sulfur amino acid homeostasis inSaccharomyces cerevisiae

  • Irène Jacquemin-Faure
  • Dominique Thomas
  • Jean Laporte
  • Christian Cibert
  • Yolande Surdin-Kerjan
Original Paper


In order to isolate new mutations impairing transcriptional regulation of sulfur metabolism inSaccharomyces cerevisiae, we used a potent genetic screen based on a gene fusion expressing XylE (fromPseudomonas putida) under the control of the promoter region ofMET25. This selection yielded strains mutated in various different genes. We describe in this paper the properties of one of them,MET27. Mutation or disruption ofMET27 leads to a methionine requirement and affects S-adenosylmethionine (AdoMet)-mediated transcriptional control of genes involved in sulfur metabolism. The cloning and sequencing ofMET27 showed that it is identical toVPS33. Disruptions or mutations of geneVPS33 are well known to impair the biogenesis and inheritance of the vacuolar compartment. However, the methionine requirement ofvps33 mutants has not been reported previously. We show here, moreover, that other vps mutants of class C (no apparent vacuoles) also require methionine for growth. Northern blotting experiments revealed that themet27-1 mutation delayed derepression of the transcription of genes involved in sulfur metabolism. By contrast, this delay was not observed in amet27 disrupted strain. Physiological and morphological analyses ofmet27-1 andmet27 disrupted strains showed that these results could be explained by alterations in the ability of the vacuole to transport or store AdoMet, the physiological effector of the transcriptional regulation of sulfur metabolism.

Key words Yeast

Sulfur metabolism Vacuolar biogenesis Transcription 


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  1. Baker RE, Masison DC (1990) Isolation of the gene encoding theSaccharomyces cerevisiae centromere binding protein CP1. Mol Cell Biol 10:2458–2467Google Scholar
  2. Baldari C, Cesareni G (1985) Plasmids pEMBLY: new single stranded shuttle vectors for the recovery and analysis of yeast DNA sequences. Gene 35:27–32Google Scholar
  3. Banta LM, Vida TA, Herman PK, Emr SD (1990) Characterization of yeast Vps33p, a protein required for vacuolar protein sorting and vacuole biogenesis. Mole Cell Biol 10:4638–4649Google Scholar
  4. Bram RJ, Kornberg RD (1987) Isolation of a centromere DNA binding protein, its homolog, and its possible role as a transcription factor. Mol Cell Biol 7:403–409Google Scholar
  5. Cai M, Davis RW (1990) Yeast centromerc binding protein CBF1, of the helix-loop-helix protein family, is required for chromosome stability and methionine prototrophy. Cell 61:437–446Google Scholar
  6. Cantoni GL (1977) S-Adensylmethionine: present status and future perspectives. In: Salvatore F, Borek E, Zappia V, Williams-Ashmann HG, Schlenk F (eds) The biochemistry of adenosylmethionine. Columbia University Press, New York, pp 557–577Google Scholar
  7. Cherest H, Surdin-Kerjan Y (1992) Genetic analysis of a new mutation conferring cysteine auxotrophy inSaccharomyces cerevisiae: updating of the sulfur metabolism pathway. Genetics 130:51–58Google Scholar
  8. Cherest H, Nguyen Ngoc T, Surdin-Kerjan Y (1985) Transcriptional regulation of theMET3 gene fromSaccharomyces cerevisiae. Gene 34:269–281Google Scholar
  9. Dang CV, Dolde C, Gillison ML, Kato GJ (1992) Discrimination between related DNA sites by a single amino acid residue of Myc related basic-helix-loop-helix proteins. Proc Natl Acad Sci USA 89:599–602Google Scholar
  10. Dessen P, Fondrat C, Valencien C, Mugnier C (1990) Bisance: a French service for access to biomolecular databases. Comput Appl Biosci 6:355–356Google Scholar
  11. Farooqui JZ, Lee HW, Kim S, Paik WK (1983) Studies on compartmentation of S-adenosyl-l-methionine inSaccharomyces cerevisiae and isolated rat hepatocytes. Biophys Biochim Acta 757:342–351Google Scholar
  12. Hieter P, Pridmore D, Hegemann JH, Thomas H, Davis RW, Philippsen P (1985) Functional selective and analysis of yeast centromeric DNA. Cell 42:913–921Google Scholar
  13. Hoffmann CS, Winston F (1987) A ten minutes DNA preparation from yeast releases autonomous plasmids for transformation ofEscherichia coli. Gene 57:267–272Google Scholar
  14. Ito H, Fukuda I, Murata K, Kimura A (1983) Transformation of intact yeast cells treated with alkali cations. J Bacteriol 153:163–168Google Scholar
  15. Jones EW (1977) Proteinase mutants ofSaccharomyces cerevisiae. Genetics 85:23–33Google Scholar
  16. Kitamoto K, Yoshizawa K, Oshumi Y, Anraku Y (1988) Mutants ofSaccharomyces cerevisiae with defective vacuolar function. Mol Cell Biol 170:2687–2691Google Scholar
  17. Klionsky DJ, Herman PK, Emr SD (1990) The fungal vacuole: composition, function and biogenesis. Microbiol Rev 54:266–292Google Scholar
  18. Köhrer K, Emr SD (1993) The yeastVPS17 gene encodes a membrane-associated protein required for the sorting of soluble vacuolar hydrolases. J Biol Chem 268:559–569Google Scholar
  19. Mellor J, Jiang W, Funk M, Rathjen J, Barnes CA, Hinz T, Hegemann JH, Philippsen P (1990) CPF1, a yeast protein which functions in centromeres and promoters. EMBO J 9:4017–4026Google Scholar
  20. Mellor J, Rathjen J, Jiang W, Dowell SJ (1991) DNA binding of CPF1 is required for optimal centromere function but not for maintaining methionine prototrophy in yeast. Nucleic Acids Res 19:2961–2969Google Scholar
  21. Ohya Y, Oshumi Y, Anraku Y (1986) Isolation and characterization of Ca++ mutants ofSaccharomyces cerevisiae. J Gen Microbiol 132:979–988Google Scholar
  22. Raymond CK, Roberts CJ, Moore KE, Howald I, Stevens TH (1992) Biogenesis of the vacuole inSaccharomyces cerevisiae. Int Rev Cytol 139:59–120Google Scholar
  23. Riva A (1974) A simple and rapid staining method for enhancing the contrast of tissues previously treated with uranyl acetate. J Microsc 19:105–108Google Scholar
  24. Salat-Trepat JM, Evans WC (1971) The meta clearvage of catechol byAzotobacter species. Eur J Biochem 20:400–413Google Scholar
  25. Schwenke J, Robichon-Szulmajster H (1976) The transport of S-adenosyl-l-methionine in isolated yeast vacuoles and spheroplasts. Eur J Biochem 65:49–60Google Scholar
  26. Sikorski RS, Hieter P (1989) A system of shuttle vectors and yeast strains designed for efficient manipulation of DNA inSaccharomyces cerevisiae. Genetics 122:19–27Google Scholar
  27. Shapiro SK, Ehninger DJ (1966) Methods for the analysis and preparation of adenosylhomocysteine. Anal Biochem 15:323–333Google Scholar
  28. Thomas PS (1980) Hybridization of denatured DNA and small DNA fragments transferred to nitrocellulose. Proc Natl Acad Sci USA 77:5201–5205Google Scholar
  29. Thomas D, Surdin-Kerjan Y (1989) Structure of theHOM2 gene ofSaccharomyces cerevisiae and regulation of its expression. Mol Gen Genet 217:149–154Google Scholar
  30. Thomas D, Surdin-Kerjan Y (1990) An improved strategy for generating a family of unidirectional deletions on large DNA fragments. Gene Anal Tech Appl 7:87–90Google Scholar
  31. Thomas D, Rothstein R, Rosenberg N, Surdin-Kerjan Y (1988) SAM2 encodes the second methionine S-adenosyl transferase inSaccharomyces cerevisiae: physiology and regulation of both enzymes. Mol Cell Biol 8:5132–5139Google Scholar
  32. Thomas D, Cherest H, Surdin-Kerjan Y (1989) Elements involved in S-adenosylmethionine mediated regulation of theSaccharomyces cerevisiaeMET25 gene. Mol Cell Biol 9:3292–3298Google Scholar
  33. Thomas D, Barbey R, Henry D, Surdin-Kerjan Y (1992a) Physiological analysis of mutants ofSaccharomyces cerevisiae impaired in sulphate assimilation. J Gen Microbiol 138:2021–2028Google Scholar
  34. Thomas D, Jacquemin I, Surdin-Kerjan Y (1992b) MET4, a leucine zipper protein, and centromere binding factor I, are both required for transcriptional activation of sulfur metabolism inSaccharomyces cerevisiae. Mol Cell Biol 12:1719–1727Google Scholar
  35. Wada Y, Kitamoto K, Kanbe T, Tanaka K, Anraku Y (1990) TheSLPI gene ofSaccharomyces cerevisiae is essential for vacuolar morphogenesis and function. Mol Cell Biol 10:2214–2223Google Scholar
  36. Wada Y, Oshumi Y, Anraku Y (1992) Genes for directing vacuolar morphogenesis inSaccharomyces cerevisiae. J Biol Chem 267:8665–8670Google Scholar
  37. Worsay MJ, Williams PA (1975) Metabolism of toluene and xylene byPseudomonas putida (arvilla) mt-2: evidence for a new function of the TOL plasmid. J Bacteriol 124:7–13Google Scholar
  38. Wright R, Rine J (1989) Transmission electron microscopy and immunocytochemical studies of yeast: analysis of HMG-CoA reductase overproduction by electron microscopy. Methods Cell Biol 31:472–512Google Scholar

Copyright information

© Springer-Verlag 1994

Authors and Affiliations

  • Irène Jacquemin-Faure
    • 1
  • Dominique Thomas
    • 1
  • Jean Laporte
    • 2
  • Christian Cibert
    • 3
  • Yolande Surdin-Kerjan
    • 1
  1. 1.Centre de Génétique MoléculaireC.N.R.S.Gif-sur-YvetteFrance
  2. 2.Laboratoire d'EnzymologieC.N.R.S.Gif-sur-YvetteFrance
  3. 3.Institut Jacques MonodParis Cedex 05France

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