1887

Abstract

Summary: strain V918 was previously isolated in a search for thermosensitive autolytic mutants and found to bear a recessive mutation that caused the development of multinucleate swollen cells undergoing cell lysis. The gene has been isolated by complementation of the phenotype of a V918 segregant. encodes a Rho-GTPase-activating protein (GAP) which is thought to act as a modulator of the Rho1 small GTPase. It is shown that the mutation causing the morphogenetic and autolytic phenotype in strain V918 and its segregants lies in the gene, defining a new mutant allele, Mutants in the gene have been reported to display loss of cell polarity and depolarization of the actin cytoskeleton, causing a bud-emergence defect. Low resistance to sonication and to hydrolytic enzymes proved that the cell wall is less protective in mutants than in wild-type strains. Moreover, mutants are more sensitive than the wild-type to several antifungal drugs. Transmission electron microscopy revealed the development of abnormally thick and wide septa and the existence of thin areas in the cell wall which probably account for cell lysis. The depolarization of actin in mutants did not preclude morphogenetic events such as cell elongation in homozygous diploid strains during nitrogen starvation in solid media, hyperpolarization of growth in a background bearing a mutated septin, or sporulation. Multinucleate cells from homozygous diploids underwent sporulation giving rise to multispored asci (‘polyads’), containing up to 36 spores. This phenomenon occurred only under osmotically stabilized conditions, suggesting that the integrity of the ascus wall is impaired in cells expressing the mutation. It is concluded that the function of the gene product is essential for the maintenance of a functional cell wall.

Loading

Article metrics loading...

/content/journal/micro/10.1099/00221287-144-1-25
1998-01-01
2024-03-28
Loading full text...

Full text loading...

/deliver/fulltext/micro/144/1/mic-144-1-25.html?itemId=/content/journal/micro/10.1099/00221287-144-1-25&mimeType=html&fmt=ahah

References

  1. Adams A., Pringle J. 1984; Relationship of actin and tubulin distribution to bud growth in wild-type and morphogenetic mutant Saccharomyces cerevisiae . J Cell Biol 98934–945
    [Google Scholar]
  2. Adams A., Johnson D. I., Longnecker R. M., Sloat B. F., Pringle J. R. 1990; CDC42 and CDC43, two additional genes involved in budding and the establishment of cell polarity in Saccharomyces cerevisiae . J Cell Biol 111131–142
    [Google Scholar]
  3. Ausubel F. M., Kingston R. E., Brent R., Moore D. D., Seidman J. G., Smith J. A., Struhl K. 1993 Current Protocols in Molecular Biology New York: Greene Publishing Associates/Wiley Interscience;
    [Google Scholar]
  4. Ballou L., Hitzeman R. A., Lewis M. S., Ballou C. E. 1991; Vanadate-resistant yeast mutants are defective in protein glycosylation. Proc Natl Acad Sci USA 883209–3212
    [Google Scholar]
  5. Bender A., Pringle J. R. 1991; Use of a screen for synthetic lethal and multicopy suppressee mutants to identify two new genes involved in morphogenesis in Saccharomyces cerevisiae . Mol Cell Biol 111295–1305
    [Google Scholar]
  6. Bénédetti H., Raths S., Crausaz F., Riezman H. 1994; The END3 gene encodes a protein that is required for the internalization step of endocytosis and for actin cytoskeleton organization in yeast. Mol Biol Cell 51023–1037
    [Google Scholar]
  7. Cabib E., Duran A. 1975; Simple and sensitive procedure for screening yeast mutants that lyse at nonpermissive temperatures. J Bacteriol 1241604–1606
    [Google Scholar]
  8. Castro C., Ribas J. C., Valdivieso M. H., Varona R., del Rey F., Durán A. 1995; Papulacandin B resistance in yeast: isolation and characterization of PBR1, a gene involved in (l,3)β-d-glucan synthesis in Saccharomyces cerevisiae . J Bacteriol 1775732–5739
    [Google Scholar]
  9. Chan C. S. M., Botstein D. 1993; Isolation and characterization of chromosome-gain and increase-in-ploidy mutants in yeast. Genetics 135677–691
    [Google Scholar]
  10. Chen G., –C., Zheng L., Chan C. S. M. 1996; The LIM domain-containing Dbm1 GTPase-activating protein is required for normal cellular morphogenesis in Saccharomyces cerevisiae . Mol Cell Biol 161376–1390
    [Google Scholar]
  11. Cid V. J., Sanchez M., Nombela C. 1994; Characterization of thermosensitive autolytic mutants from diploid Saccharomyces cerevisiae . Microbiology 140559–568
    [Google Scholar]
  12. Cid V. J., Durán A., del Rey F., Snyder M., Nombela C., Sánchez M. 1995; Molecular basis of cell integrity and morphogenesis in Saccharomyces cerevisiae . Microbiol Rev 59345–386
    [Google Scholar]
  13. Donnelly S. F. H., Pocklington M. J., Pallotta D., Orr E. 1993; A proline-rich protein, verprolin, involved in cytoskeletal organization and cellular growth in the yeast Saccharomyces cerevisiae . Mol Microbiol 10585–596
    [Google Scholar]
  14. Douglas C. M., Marrinan W. L., Li W., Kurtz M. B. 1994; A Saccharomyces cerevisiae mutant with echinocandin-resistant1,3-β-d-glucan synthase. J Bacteriol 1765686–5696
    [Google Scholar]
  15. Drgonová J., Drgon T., Tanaka K., Kollár R., Chen G. -C., Ford R. A., Chan C. S. M., Takai Y., Cabib E. 1996; Rho1p, a yeast protein at the interface between cell polarization and morphogenesis. Science 272277–279
    [Google Scholar]
  16. Elledge S. J., Davis R. W. 1988; A family of versatile centromeric vectors designed for use in the sectoring-shuffle mutagenesis assay in Saccharomyces cerevisiae. . Gene 70303–312
    [Google Scholar]
  17. Gabriel M., Kopecká M. 1995; Disruption of the actin cytoskeleton in budding yeast results in formation of an aberrant cell wall. Microbiology 141891–899
    [Google Scholar]
  18. Gimeno C. J., Ljungdahl P. O., Styles C. A., Fink G. R. 1992; Unipolar cell divisions in the yeast Saccharomyces cerevisiae lead to filamentous growth: regulation by starvation and RAS. . Cell 681077–1090
    [Google Scholar]
  19. Hartwell L. H. 1971; Genetic control of cell division cycle in yeast. IV. Genes controlling bud emergence and cytokinesis. Exp Cell Res 69265–276
    [Google Scholar]
  20. Healy A. M., Zolnierowicz S., Stapleton A. E., Goebl M., DePaoli-Roach A. A., Pringle J. R. 1991; CDC55, a Saccharomyces cerevisiae gene involved in cellular morphogenesis: identification, characterization, and homology to the B subunit of mammalian type 2A protein phosphatase. Mol Cell Biol 115767–5780
    [Google Scholar]
  21. Johnston G. C., Prendergast J. A., Singer R. A. 1991; The Saccharomyces cerevisiae MYO2 gene encodes an essential myosin for vectorial transport of vesicles. J Cell Biol 113539–551
    [Google Scholar]
  22. Kim Y. -J., Francisco L., Chen G. -C., Marcotte E., Chan C. S. M. 1994; Control of cellular morphogenesis by the Ipl2/Bem2 GTPase-activating protein: possible role of protein phosphorylation. J Cell Biol 1271381–1384
    [Google Scholar]
  23. Klis F. M. 1994; Cell wall assembly in yeast. Yeast 10851–869
    [Google Scholar]
  24. Kohno H., Tanaka K., Mino A., Umikawa M., Imamura H., Fujiwara T., Fujita Y., Hotta K., Qadota H., Watanabe T., Ohya Y., Takai Y. 1996; Bni1p implicated in cytoskeletal control is a putative target of Rho1p small GTP binding protein in Saccharomyces cerevisiae. . EMBO J 156060–6068
    [Google Scholar]
  25. Kuo S. C., Lampen J. O. 1974; Tunicamycin – an inhibitor of yeast glycoprotein synthesis. Biochem Biophys Res Commun 58287–295
    [Google Scholar]
  26. Lee K. S., Irie K., Gotoh Y., Watanabe Y., Araki H., Nishida E., Matsumoto K., Levin D. E. 1993; A yeast mitogen-activated protein kinase homolog (Mpk1p) mediates signalling by protein kinase C. Mol Cell Biol 133067–3075
    [Google Scholar]
  27. Liu H., Bretscher A. 1992; Characterization of TPM1 disrupted yeast cells indicates an involvement of tropomyosin in directed vesicular transport. J Cell Biol 118285–299
    [Google Scholar]
  28. Longtine M. S., DeMarini D., Valencik M. L., Al-Awar O. S., Fares H., De Virgilio C., Pringle J. R. 1996; The septins: roles in cytokinesis and other processes. Curr Opin Cell Biol 8106–119
    [Google Scholar]
  29. Madaule P., Axel R., Myers A. M. 1987; Characterization of two members of the rho gene family from the yeast Saccharomyces cerevisiae . Proc Natl Acad Sci USA 84779–783
    [Google Scholar]
  30. Martin H., Arroyo J., Sanchez M., Molina M., Nombela C. 1993; Activity of the yeast MAP kinase homologue Slt2 is critically required for cell integrity at 37°C. Mol Gen Genet 241177–184
    [Google Scholar]
  31. Masuda T., Tanaka K., Nonaka H., Yamochi W., Maeda A., Takai Y. 1994; Molecular cloning and characterization of yeast Rho GDP dissociation inhibitor. J Biol Chem 26919713–19718
    [Google Scholar]
  32. Miret J. J., Solari A. J., Barderi P. A., Goldemberg S. H. 1992; Polyamines and cell wall organization in Saccharomyces cerevisiae . Yeast 81033–1041
    [Google Scholar]
  33. Mulholland J., Preuss D., Moon A., Wong A., Drubin D., Botstein D. 1994; Ultrastructure of the yeast actin cytoskeleton and its association with the plasma membrane. J Cell Biol 125381–391
    [Google Scholar]
  34. Nonaka H., Tanaka K., Hirano H., Fujiwara T., Kohno H., Umikawa M., Mino A., Takai Y. 1995; A downstream target of RHO1 small GTP-binding protein is PKC1, a homolog of protein kinase C, which leads to activation of the MAP kinase cascade in Saccharomyces cerevisiae . EMBO J 145931–5938
    [Google Scholar]
  35. Ozaki K., Tanaka K., Imamura H., Hiraha T., Kameyama T., Nonaka H., Hirano H., Matsuura Y., Takai T. 1996; Rom1p and Rom2p are GDP/GTP exchange proteins (GEPs) for the Rho1p small GTP binding protein in Saccharomyces cerevisiae . EMBO J 152196–2207
    [Google Scholar]
  36. Paravicini G., Cooper M., Friedli L., Smith D. J., Carpentier J. L., Klig L. S., Payton M. A. 1992; The osmotic integrity of the yeast cell requires a functional PKC1 gene product. Mol Cell Biol 124896–4905
    [Google Scholar]
  37. Peterson J., Zheng Y., Bender, L, Myers A., Cerione R., Bender A. 1994; Interactions between the bud emergence proteins Bem1p and Bem2p and Rho-type GTPases in yeast. J Cell Biol 1271395–1406
    [Google Scholar]
  38. Qadota H., Python C. P., Inoue S. B., Arisawa M., Anraku Y., Zheng Y., Watanabe T., Levin D. E., Ohya Y. 1996; Identification of yeast Rho1p GTPase as a regulatory subunit of 1,3-beta-glucan synthase. Science 272279–281
    [Google Scholar]
  39. Ridley A. J. 1995; Rho-related proteins: actin cytoskeleton and cell cycle. Curr Opin Genet Dev 524–30
    [Google Scholar]
  40. Sambrook J., Fritsch E. F., Maniatis T. 1989 Molecular Cloning: a Laboratory Manual 2nd edn Cold Spring Harbor, NY: Cold Spring Harbor Laboratory;
    [Google Scholar]
  41. Schmidt A., Bickle M., Beck T., Hall M. N. 1997; The yeast phosphatidylinositol kinase homolog TOR2 activates RHO1 and RHO2 via the exchange factor ROM2 . Cell 88531–542
    [Google Scholar]
  42. Sikorski R. S., Hieter P. 1989; A system of shuttle vectors and yeast host strains designed for efficient manipulation of DNA in Saccharomyces cerevisiae . Genetics 12219–27
    [Google Scholar]
  43. Sloat B. F., Adams A., Pringle J. R. 1981; Roles of the CDC24 gene product in cellular morphogenesis during the Saccharomyces cerevisiae cell cycle. J Cell Biol 89395–405
    [Google Scholar]
  44. Stevenson B. J., Ferguson B., De Virgilio C., Bi E., Pringle J. R., Ammerer G., Sprague G. F. Jr 1995; Mutation of RGA1 which encodes a putative GTPase-activating protein for the polarity-establishment protein Cdc42p, activates the pheromone-response pathway in the yeast Saccharomyces cerevisiae . Genes Dev 92949–2963
    [Google Scholar]
  45. Torres L., Martin H., Garcia-Saez M. I., Arroyo J., Molina M., Sanchez M., Nombela C. 1991; A protein kinase gene complements the lytic phenotype of Saccharomyces cerevisiae lyt2 mutants. Mol Microbiol 52845–2854
    [Google Scholar]
  46. Wang T., Bretscher A. 1995; The rho-GAP encoded by BEM2 regulates cytoskeletal structure in budding yeast. Mol Biol Cell 61011–1024
    [Google Scholar]
  47. Yamochi W., Tanaka K., Nonaka H., Maeda A., Musha T., Takai Y. 1994; Growth site localization of Rho1 small GTP-binding protein and its involvement in bud formation in Saccharomyces cerevisiae . J Cell Biol 1251077–1093
    [Google Scholar]
  48. Zarzov P., Mazzoni C., Mann C. 1996; The SLT2(MPK1) MAP kinase is activated during periods of polarized cell growth in yeast.
  49. Zheng Y., Cerione R., Bender A. 1994; Control of the yeast bud-site assembly GTPase Cdc42. J Biol Chem 2692369–2372
    [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/micro/10.1099/00221287-144-1-25
Loading
/content/journal/micro/10.1099/00221287-144-1-25
Loading

Data & Media loading...

This is a required field
Please enter a valid email address
Approval was a Success
Invalid data
An Error Occurred
Approval was partially successful, following selected items could not be processed due to error