Molecular Genetics and Genomics

, Volume 284, Issue 3, pp 217–229 | Cite as

A block of endocytosis of the yeast cell wall integrity sensors Wsc1 and Wsc2 results in reduced fitness in vivo

  • Sabrina Wilk
  • Janina Wittland
  • Andreas Thywissen
  • Hans-Peter Schmitz
  • Jürgen J. HeinischEmail author
Original Paper


The response to cell surface stress in yeast is mediated by a set of five plasma membrane sensors. We here address the relation of intracellular localization of the sensors Wsc1, Wsc2, and Mid2 to their turnover and signaling function. Growth competition experiments indicate that Wsc2 plays an important role in addition to Wsc1 and Mid2. The two Wsc sensors appear at the bud neck during cytokinesis and employ different routes of endocytosis, which govern their turnover. Whereas Wsc1 uses a clathrin-dependent NPFDD signal, Wsc2 relies on a specific lysine residue (K495). In end3 and doa4 endocytosis mutants, both sensors accumulate at the plasma membrane, and a hypersensitivity to cell wall-specific drugs and to treatment with zymolyase is observed. A haploid strain in which endocytosis of the two sensors is specifically blocked displays a reduced fitness in growth competition experiments. If the Mid2 sensor is mobilized by the addition of an endocytosis signal, it mimics the dynamic distribution of the Wsc sensors, but is unable to complement the specific growth defects of a wsc1 deletion. These data suggest that sensor distribution is not the major determinant for its specificity.


CWI signaling Wsc1/Slg1 Wsc2 Mid2 Endocytosis 



We are grateful to Arne Jendretzki for mCherry plasmids and the Myo1-fusion strain, and to Britta Delvos for the strain carrying an in-frame deletion within the MID2 sequence encoding the cytoplasmic tail. We also would like to thank Bernadette Sander-Turgut and Eugenia Leno for technical assistance and Rosaura Rodicio for very critical reading of the manuscript and intensive discussions. This work was made possible by a grant from the Deutsche Forschungsgemeinschaft (SFB431).

Supplementary material

438_2010_563_MOESM1_ESM.pdf (732 kb)
Supplementary material 1 (PDF 732 kb)


  1. Abe M, Qadota H, Hirata A, Ohya Y (2003) Lack of GTP-bound Rho1p in secretory vesicles of Saccharomyces cerevisiae. J Cell Biol 162:85–97CrossRefPubMedGoogle Scholar
  2. Alonso-Monge R, Real E, Wojda I, Bebelman JP, Mager WH, Siderius M (2001) Hyperosmotic stress response and regulation of cell wall integrity in Saccharomyces cerevisiae share common functional aspects. Mol Microbiol 41:717–730CrossRefPubMedGoogle Scholar
  3. Arvanitidis A, Heinisch JJ (1994) Studies on the function of yeast phosphofructokinase subunits by in vitro mutagenesis. J Biol Chem 269:8911–8918PubMedGoogle Scholar
  4. Conant GC, Wolfe KH (2007) Increased glycolytic flux as an outcome of whole-genome duplication in yeast. Mol Syst Biol 3:129CrossRefPubMedGoogle Scholar
  5. Delley PA, Hall MN (1999) Cell wall stress depolarizes cell growth via hyperactivation of Rho1. J Cell Biol 147:163–174CrossRefPubMedGoogle Scholar
  6. Dupres V, Alsteens D, Wilk S, Hansen B, Heinisch JJ, Dufrene YF (2009) The yeast Wsc1 cell surface sensor behaves like a nanospring in vivo. Nat Chem Biol 5:857–862CrossRefPubMedGoogle Scholar
  7. Galan JM, Peter M (1999) Ubiquitin-dependent degradation of multiple F-box proteins by an autocatalytic mechanism. Proc Natl Acad Sci USA 96:9124–9129CrossRefPubMedGoogle Scholar
  8. Gietz RD, Sugino A (1988) New yeast-Escherichia coli shuttle vectors constructed with in vitro mutagenized yeast genes lacking six-base pair restriction sites. Gene 74:527–534CrossRefPubMedGoogle Scholar
  9. Green R, Lesage G, Sdicu AM, Menard P, Bussey H (2003) A synthetic analysis of the Saccharomyces cerevisiae stress sensor Mid2p, and identification of a Mid2p-interacting protein, Zeo1p, that modulates the PKC1-MPK1 cell integrity pathway. Microbiology 149:2487–2499CrossRefPubMedGoogle Scholar
  10. Hasegawa Y, Irie K, Gerber AP (2008) Distinct roles for Khd1p in the localization and expression of bud-localized mRNAs in yeast. RNA 14:2333–2347CrossRefPubMedGoogle Scholar
  11. Heinisch JJ (2008) Baker’s yeast as a tool for the development of antifungal drugs which target cell integrity—an update. Expert Opin Drug Discov 3:931–943CrossRefGoogle Scholar
  12. Heinisch JJ, Lorberg A, Schmitz HP, Jacoby JJ (1999) The protein kinase C-mediated MAP kinase pathway involved in the maintenance of cellular integrity in Saccharomyces cerevisiae. Mol Microbiol 32:671–680CrossRefPubMedGoogle Scholar
  13. Heinisch JJ, Dupres V, Wilk S, Jendretzki A, Dufrene YF (2010) Single-molecule atomic force microscopy reveals clustering of the yeast plasma-membrane sensor Wsc1. PLoS One 5(6):e11104Google Scholar
  14. Hicke L, Riezman H (1996) Ubiquitination of a yeast plasma membrane receptor signals its ligand-stimulated endocytosis. Cell 84:277–287CrossRefPubMedGoogle Scholar
  15. Hutzler F, Gerstl R, Lommel M, Strahl S (2008) Protein N-glycosylation determines functionality of the Saccharomyces cerevisiae cell wall integrity sensor Mid2p. Mol Microbiol 68:1438–1449CrossRefPubMedGoogle Scholar
  16. Jendretzki A, Ciklic I, Rodicio R, Schmitz HP, Heinisch JJ (2009) Cyk3 acts in actomyosin ring independent cytokinesis by recruiting Inn1 to the yeast bud neck. Mol Genet Genomics 282:437–451CrossRefPubMedGoogle Scholar
  17. Ketela T, Green R, Bussey H (1999) Saccharomyces cerevisiae Mid2p is a potential cell wall stress sensor and upstream activator of the PKC1-MPK1 cell integrity pathway. J Bacteriol 181:3330–3340PubMedGoogle Scholar
  18. Kirchrath L, Lorberg A, Schmitz HP, Gengenbacher U, Heinisch JJ (2000) Comparative genetic and physiological studies of the MAP kinase Mpk1p from Kluyveromyces lactis and Saccharomyces cerevisiae. J Mol Biol 300:743–758CrossRefPubMedGoogle Scholar
  19. Klis FM, Mol P, Hellingwerf K, Brul S (2002) Dynamics of cell wall structure in Saccharomyces cerevisiae. FEMS Microbiol Rev 26:239–256CrossRefPubMedGoogle Scholar
  20. Levin DE (2005) Cell wall integrity signaling in Saccharomyces cerevisiae. Microbiol Mol Biol Rev 69:262–291CrossRefPubMedGoogle Scholar
  21. Lodder AL, Lee TK, Ballester R (1999) Characterization of the Wsc1 protein, a putative receptor in the stress response of Saccharomyces cerevisiae. Genetics 152:1487–1499PubMedGoogle Scholar
  22. Lommel M, Bagnat M, Strahl S (2004) Aberrant processing of the WSC family and Mid2p cell surface sensors results in cell death of Saccharomyces cerevisiae O-mannosylation mutants. Mol Cell Biol 24:46–57CrossRefPubMedGoogle Scholar
  23. Philip B, Levin DE (2001) Wsc1 and Mid2 are cell surface sensors for cell wall integrity signaling that act through Rom2, a guanine nucleotide exchange factor for Rho1. Mol Cell Biol 21:271–280CrossRefPubMedGoogle Scholar
  24. Piao HL, Machado IM, Payne GS (2007) NPFXD-mediated endocytosis is required for polarity and function of a yeast cell wall stress sensor. Mol Biol Cell 18:57–65CrossRefPubMedGoogle Scholar
  25. Rajavel M, Philip B, Buehrer BM, Errede B, Levin DE (1999) Mid2 is a putative sensor for cell integrity signaling in Saccharomyces cerevisiae. Mol Cell Biol 19:3969–3976PubMedGoogle Scholar
  26. Rodicio R, Buchwald U, Schmitz HP, Heinisch JJ (2008) Dissecting sensor functions in cell wall integrity signaling in Kluyveromyces lactis. Fungal Genet Biol 45:422–435CrossRefPubMedGoogle Scholar
  27. Schmidt M, Bowers B, Varma A, Roh DH, Cabib E (2002) In budding yeast, contraction of the actomyosin ring and formation of the primary septum at cytokinesis depend on each other. J Cell Sci 115:293–302PubMedGoogle Scholar
  28. Straede A, Heinisch JJ (2007) Functional analyses of the extra- and intracellular domains of the yeast cell wall integrity sensors Mid2 and Wsc1. FEBS Lett 581:4495–4500CrossRefPubMedGoogle Scholar
  29. Tan PK, Howard JP, Payne GS (1996) The sequence NPFXD defines a new class of endocytosis signal in Saccharomyces cerevisiae. J Cell Biol 135:1789–1800CrossRefPubMedGoogle Scholar
  30. Tang HY, Xu J, Cai M (2000) Pan1p, End3p, and S1a1p, three yeast proteins required for normal cortical actin cytoskeleton organization, associate with each other and play essential roles in cell wall morphogenesis. Mol Cell Biol 20:12–25CrossRefPubMedGoogle Scholar
  31. Valdez-Taubas J, Pelham HR (2003) Slow diffusion of proteins in the yeast plasma membrane allows polarity to be maintained by endocytic cycling. Curr Biol 13:1636–1640CrossRefPubMedGoogle Scholar
  32. Verna J, Lodder A, Lee K, Vagts A, Ballester R (1997) A family of genes required for maintenance of cell wall integrity and for the stress response in Saccharomyces cerevisiae. Proc Natl Acad Sci USA 94:13804–13809CrossRefPubMedGoogle Scholar

Copyright information

© Springer-Verlag 2010

Authors and Affiliations

  • Sabrina Wilk
    • 1
  • Janina Wittland
    • 1
  • Andreas Thywissen
    • 1
  • Hans-Peter Schmitz
    • 1
  • Jürgen J. Heinisch
    • 1
    Email author
  1. 1.AG Genetik, Fachbereich Biologie/ChemieUniversität OsnabrückOsnabrückGermany

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