1887

Abstract

The yeast cell wall contains an unusually high number of different mannoproteins. The physiological role of most of them is unknown and gene disruptions leading to depletion of different proteins do not affect major functions of the wall. In this work the phenotype of different single and multiple cell wall protein mutants was observed at the level of individual cells. It was found that the lack of the non-covalently bound wall proteins Scw4p, Scw10p and Bgl2p increases the mortality of cells grown exponentially under standard laboratory conditions, as assayed by methylene blue staining. Mutation of , however, suppressed the phenotype of , or , indicating that Scw4p, Scw10p and Bgl2p act synergistically while Scw11p has an activity antagonistic to that of the other three proteins. Mutants lacking major covalently bound proteins, either all four described Pir-proteins or the five most abundant glycosylphosphatidylinositol (GPI)-anchored proteins (Ccw12p, Ccw13p/Dan1p, Ccw14p/Icwp1p, Tip1p and Cwp1p), also had increased mortalities, the first somewhat more and the latter less than that of . In all cases the observed phenotype was suppressed by the addition of an osmotic stabilizer to the growth medium, indicating that cells died due to decreased osmotic stability. If cells were grown to stationary phase, Scw-mutants showed only slightly increased mortality, but mutants lacking Pir- or GPI-anchored proteins had significantly increased sensitivity, suggesting that their physiological function is primarily expressed in stationary-phase cells. In many cases structures consisting of a living (multiple Pir-protein mutant) mother with two methylene blue-stained daughters could be seen.

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2004-10-01
2024-04-20
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References

  1. Cappellaro, C., Mrša, V. & Tanner, W.(1998). New potential cell wall glucanases of Saccharomyces cerevisiae and their involvement in mating. J Bacteriol 180, 5030–5037. [Google Scholar]
  2. Eamus, D. G. & Jennings, D. H.(1986). Water, turgor and osmotic potentials of fungi. In Water, Fungi and Plants, pp. 27–47. Edited by P. G. Ayres & L. Body. Cambridge: Cambridge University Press.
  3. Fleet, G. H.(1991). Cell walls. In The Yeasts, 2nd edn, pp. 199 – 277. Edited by A. H. Rose & G. H. Harrison. London: Academic Press.
  4. Garlier, D. & Thomasett, N.(1986). Use of MTT colorimetric assay to measure cell activation. J Immunol Methods 94, 57–63.[CrossRef] [Google Scholar]
  5. Klebl, F. & Tanner, W.(1989). Molecular cloning of a cell wall exo-β-1,3-glucanase from Saccharomyces cerevisiae. J Bacteriol 171, 6259–6264. [Google Scholar]
  6. Klis, F. M.(1994). Review: cell wall assembly in yeast. Yeast 10, 851–869.[CrossRef] [Google Scholar]
  7. Montijn, R. C., van Rinsum, J., Van Schagen, F. A. & Klis, F. M.(1994). Glucomannoproteins in the cell wall of Saccharomyces cerevisiae contain a novel type of carbohydrate side-chain. J Biol Chem 269, 19338–19342. [Google Scholar]
  8. Moukadiri, I., Armero, J., Abad, A., Sentandreu, R. & Zueco, J.(1997). Identification of a mannoprotein present in the inner layer of the cell wall of Saccharomyces cerevisiae. J Bacteriol 179, 2154–2162. [Google Scholar]
  9. Mouyna, I., Fontaine, T., Vai, M., Monod, M., Fonzi, W. A., Diaquin, M., Popolo, L., Hartland, R. P. & Latge, J.-P.(2000). Glycosylphosphatidylinositol-anchored glucanosyltransferases play an active role in the biosynthesis of the fungal cell wall. J Biol Chem 275, 14882–14889.[CrossRef] [Google Scholar]
  10. Mrša, V. & Tanner, W.(1999). Role of NaOH-extractable cell wall proteins Ccw5p, Ccw6p, Ccw7p and Ccw8p (members of the Pir protein family) in stability of the Saccharomyces cerevisiae cell wall. Yeast 15, 813–820.[CrossRef] [Google Scholar]
  11. Mrša, V., Klebl, F. & Tanner, W.(1993). Purification and characterisation of the Saccharomyces cerevisiae BGL2 gene product, a cell wall endo-β-1,3-glucanase. J Bacteriol 175, 2102–2106. [Google Scholar]
  12. Mrša, V., Seidl, T., Gentzsch, M. & Tanner, W.(1997). Specific labelling of cell wall proteins by biotinylation. Identification of four covalently linked O-mannosylated proteins of Saccharomyces cerevisiae. Yeast 13, 1145–1154.[CrossRef] [Google Scholar]
  13. Mrša, V., Ecker, M., Strahl-Bolsinger, S., Nimtz, M., Lehle, L. & Tanner, W.(1999). Deletion of new covalently linked cell wall glycoproteins alters the electrophoretic mobility of phosphorylated wall components of Saccharomyces cerevisiae. J Bacteriol 181, 3076–3086. [Google Scholar]
  14. Popolo, L. & Vai, M.(1999). The Gas1 glycoprotein, a putative polymer cross-linker. Biochem Biophys Acta 1426, 385–400.[CrossRef] [Google Scholar]
  15. Robinson, J. S., Klionsky, D. J., Banta, L. M. & Emr, S. D.(1988). Protein sorting in Saccharomyces cerevisiae: isolation of mutants defective in the delivery and processing of multiple vacuolar hydrolases. Mol Cell Biol 8, 4936–4948. [Google Scholar]
  16. Rodriguez-Pena, J. M., Cid, V. J., Arroyo, J. & Nombela, C.(2000). A novel family of cell wall-related proteins regulated differently during the yeast life cycle. Mol Cell Biol 20, 3245–3255.[CrossRef] [Google Scholar]
  17. Smart, K., Chambers, K., Lambert, I. & Jenkins, C.(1999). Use of methylene violet staining procedures to determine yeast viability and vitality. J Am Soc Brew Chem 57, 18–23. [Google Scholar]
  18. Terashima, H., Hamada, K. & Kitada, K.(2003). The localisation change of Ybr078w/Ecm33, a yeast GPI-associated protein, from the plasma membrane to the cell wall, affecting the cellular function. FEMS Microbiol Lett 218, 175–180.[CrossRef] [Google Scholar]
  19. Valentin, E., Herrero, W., Pastor, J. F. I. & Sentandreu, R.(1984). Solubilization and analysis of mannoprotein molecules from the cell wall of Saccharomyces cerevisiae. J Gen Microbiol 130, 1419–1428. [Google Scholar]
  20. Van Der Vaart, J. M., Caro, L. H. P., Chapman, J. W., Klis, F. M. & Verrips, C. T.(1995). Identification of three mannoproteins in the cell wall of Saccharomyces cerevisiae. J Bacteriol 177, 3104–3110. [Google Scholar]
  21. Zheng, Y. T. & Ben, K. L.(1992). Use of MTT assay for the determination of cell viability and proliferation. Immunol J 8, 266–269. [Google Scholar]
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