Skip to main content
PMC home page
Genetics logoLink to Genetics
. 2004 Jan;166(1):67–77. doi: 10.1534/genetics.166.1.67

Fus1p interacts with components of the Hog1p mitogen-activated protein kinase and Cdc42p morphogenesis signaling pathways to control cell fusion during yeast mating.

Bryce Nelson 1, Ainslie B Parsons 1, Marie Evangelista 1, Karen Schaefer 1, Kathy Kennedy 1, Steven Ritchie 1, Tracey L Petryshen 1, Charles Boone 1
PMCID: PMC1470681  PMID: 15020407

Abstract

Cell fusion in the budding yeast Saccharomyces cerevisiae is a temporally and spatially regulated process that involves degradation of the septum, which is composed of cell wall material, and occurs between conjugating cells within a prezygote, followed by plasma membrane fusion. The plasma membrane protein Fus1p is known to be required for septum degradation during cell fusion, yet its role at the molecular level is not understood. We identified Sho1p, an osmosensor for the HOG MAPK pathway, as a binding partner for Fus1 in a two-hybrid screen. The Sho1p-Fus1p interaction occurs directly and is mediated through the Sho1p-SH3 domain and a proline-rich peptide ligand on the Fus1p COOH-terminal cytoplasmic region. The cell fusion defect associated with fus1Delta mutants is suppressed by a sho1Delta deletion allele, suggesting that Fus1p negatively regulates Sho1p signaling to ensure efficient cell fusion. A two-hybrid matrix containing fusion proteins and pheromone response pathway signaling molecules reveals that Fus1p may participate in a complex network of interactions. In particular, the Fus1p cytoplasmic domain interacts with Chs5p, a protein required for secretion of specialized Chs3p-containing vesicles during bud development, and chs5Delta mutants were defective in cell surface localization of Fus1p. The Fus1p cytoplasmic domain also interacts with the activated GTP-bound form of Cdc42p and the Fus1p-SH3 domain interacts with Bni1p, a yeast formin that participates in cell fusion and controls the assembly of actin cables to polarize secretion in response to Cdc42p signaling. Taken together, our results suggest that Fus1p acts as a scaffold for the assembly of a cell surface complex involved in polarized secretion of septum-degrading enzymes and inhibition of HOG pathway signaling to promote cell fusion.

Full Text

The Full Text of this article is available as a PDF (391.8 KB).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Adamo J. E., Moskow J. J., Gladfelter A. S., Viterbo D., Lew D. J., Brennwald P. J. Yeast Cdc42 functions at a late step in exocytosis, specifically during polarized growth of the emerging bud. J Cell Biol. 2001 Nov 12;155(4):581–592. doi: 10.1083/jcb.200106065. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Amberg D. C., Zahner J. E., Mulholland J. W., Pringle J. R., Botstein D. Aip3p/Bud6p, a yeast actin-interacting protein that is involved in morphogenesis and the selection of bipolar budding sites. Mol Biol Cell. 1997 Apr;8(4):729–753. doi: 10.1091/mbc.8.4.729. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Amberg D. C., Zahner J. E., Mulholland J. W., Pringle J. R., Botstein D. Aip3p/Bud6p, a yeast actin-interacting protein that is involved in morphogenesis and the selection of bipolar budding sites. Mol Biol Cell. 1997 Apr;8(4):729–753. doi: 10.1091/mbc.8.4.729. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Brizzio V., Gammie A. E., Nijbroek G., Michaelis S., Rose M. D. Cell fusion during yeast mating requires high levels of a-factor mating pheromone. J Cell Biol. 1996 Dec;135(6 Pt 2):1727–1739. doi: 10.1083/jcb.135.6.1727. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Brizzio V., Gammie A. E., Rose M. D. Rvs161p interacts with Fus2p to promote cell fusion in Saccharomyces cerevisiae. J Cell Biol. 1998 May 4;141(3):567–584. doi: 10.1083/jcb.141.3.567. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Buehrer B. M., Errede B. Coordination of the mating and cell integrity mitogen-activated protein kinase pathways in Saccharomyces cerevisiae. Mol Cell Biol. 1997 Nov;17(11):6517–6525. doi: 10.1128/mcb.17.11.6517. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Butty A. C., Pryciak P. M., Huang L. S., Herskowitz I., Peter M. The role of Far1p in linking the heterotrimeric G protein to polarity establishment proteins during yeast mating. Science. 1998 Nov 20;282(5393):1511–1516. doi: 10.1126/science.282.5393.1511. [DOI] [PubMed] [Google Scholar]
  8. Chenevert J., Valtz N., Herskowitz I. Identification of genes required for normal pheromone-induced cell polarization in Saccharomyces cerevisiae. Genetics. 1994 Apr;136(4):1287–1296. doi: 10.1093/genetics/136.4.1287. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Dorer R., Boone C., Kimbrough T., Kim J., Hartwell L. H. Genetic analysis of default mating behavior in Saccharomyces cerevisiae. Genetics. 1997 May;146(1):39–55. doi: 10.1093/genetics/146.1.39. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Durfee T., Becherer K., Chen P. L., Yeh S. H., Yang Y., Kilburn A. E., Lee W. H., Elledge S. J. The retinoblastoma protein associates with the protein phosphatase type 1 catalytic subunit. Genes Dev. 1993 Apr;7(4):555–569. doi: 10.1101/gad.7.4.555. [DOI] [PubMed] [Google Scholar]
  11. Elion E. A., Grisafi P. L., Fink G. R. FUS3 encodes a cdc2+/CDC28-related kinase required for the transition from mitosis into conjugation. Cell. 1990 Feb 23;60(4):649–664. doi: 10.1016/0092-8674(90)90668-5. [DOI] [PubMed] [Google Scholar]
  12. Elion E. A. Pheromone response, mating and cell biology. Curr Opin Microbiol. 2000 Dec;3(6):573–581. doi: 10.1016/s1369-5274(00)00143-0. [DOI] [PubMed] [Google Scholar]
  13. Elion E. A., Trueheart J., Fink G. R. Fus2 localizes near the site of cell fusion and is required for both cell fusion and nuclear alignment during zygote formation. J Cell Biol. 1995 Sep;130(6):1283–1296. doi: 10.1083/jcb.130.6.1283. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Evangelista M., Blundell K., Longtine M. S., Chow C. J., Adames N., Pringle J. R., Peter M., Boone C. Bni1p, a yeast formin linking cdc42p and the actin cytoskeleton during polarized morphogenesis. Science. 1997 Apr 4;276(5309):118–122. doi: 10.1126/science.276.5309.118. [DOI] [PubMed] [Google Scholar]
  15. Evangelista Marie, Pruyne David, Amberg David C., Boone Charles, Bretscher Aanthony. Formins direct Arp2/3-independent actin filament assembly to polarize cell growth in yeast. Nat Cell Biol. 2002 Jan;4(1):32–41. doi: 10.1038/ncb718. [DOI] [PubMed] [Google Scholar]
  16. Fujimura H. Molecular cloning of the DAC2/FUS3 gene essential for pheromone-induced G1-arrest of the cell cycle in Saccharomyces cerevisiae. Curr Genet. 1990 Dec;18(5):395–400. doi: 10.1007/BF00309907. [DOI] [PubMed] [Google Scholar]
  17. Fujiwara T., Tanaka K., Mino A., Kikyo M., Takahashi K., Shimizu K., Takai Y. Rho1p-Bni1p-Spa2p interactions: implication in localization of Bni1p at the bud site and regulation of the actin cytoskeleton in Saccharomyces cerevisiae. Mol Biol Cell. 1998 May;9(5):1221–1233. doi: 10.1091/mbc.9.5.1221. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Gammie A. E., Brizzio V., Rose M. D. Distinct morphological phenotypes of cell fusion mutants. Mol Biol Cell. 1998 Jun;9(6):1395–1410. doi: 10.1091/mbc.9.6.1395. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Gyuris J., Golemis E., Chertkov H., Brent R. Cdi1, a human G1 and S phase protein phosphatase that associates with Cdk2. Cell. 1993 Nov 19;75(4):791–803. doi: 10.1016/0092-8674(93)90498-f. [DOI] [PubMed] [Google Scholar]
  20. Hagen D. C., McCaffrey G., Sprague G. F., Jr Pheromone response elements are necessary and sufficient for basal and pheromone-induced transcription of the FUS1 gene of Saccharomyces cerevisiae. Mol Cell Biol. 1991 Jun;11(6):2952–2961. doi: 10.1128/mcb.11.6.2952. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Heiman M. G., Walter P. Prm1p, a pheromone-regulated multispanning membrane protein, facilitates plasma membrane fusion during yeast mating. J Cell Biol. 2000 Oct 30;151(3):719–730. doi: 10.1083/jcb.151.3.719. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Hollenberg S. M., Sternglanz R., Cheng P. F., Weintraub H. Identification of a new family of tissue-specific basic helix-loop-helix proteins with a two-hybrid system. Mol Cell Biol. 1995 Jul;15(7):3813–3822. doi: 10.1128/mcb.15.7.3813. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. James P., Halladay J., Craig E. A. Genomic libraries and a host strain designed for highly efficient two-hybrid selection in yeast. Genetics. 1996 Dec;144(4):1425–1436. doi: 10.1093/genetics/144.4.1425. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Longtine M. S., McKenzie A., 3rd, Demarini D. J., Shah N. G., Wach A., Brachat A., Philippsen P., Pringle J. R. Additional modules for versatile and economical PCR-based gene deletion and modification in Saccharomyces cerevisiae. Yeast. 1998 Jul;14(10):953–961. doi: 10.1002/(SICI)1097-0061(199807)14:10<953::AID-YEA293>3.0.CO;2-U. [DOI] [PubMed] [Google Scholar]
  25. Longtine M. S., McKenzie A., 3rd, Demarini D. J., Shah N. G., Wach A., Brachat A., Philippsen P., Pringle J. R. Additional modules for versatile and economical PCR-based gene deletion and modification in Saccharomyces cerevisiae. Yeast. 1998 Jul;14(10):953–961. doi: 10.1002/(SICI)1097-0061(199807)14:10<953::AID-YEA293>3.0.CO;2-U. [DOI] [PubMed] [Google Scholar]
  26. Maeda T., Takekawa M., Saito H. Activation of yeast PBS2 MAPKK by MAPKKKs or by binding of an SH3-containing osmosensor. Science. 1995 Jul 28;269(5223):554–558. doi: 10.1126/science.7624781. [DOI] [PubMed] [Google Scholar]
  27. McCaffrey G., Clay F. J., Kelsay K., Sprague G. F., Jr Identification and regulation of a gene required for cell fusion during mating of the yeast Saccharomyces cerevisiae. Mol Cell Biol. 1987 Aug;7(8):2680–2690. doi: 10.1128/mcb.7.8.2680. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. O'Rourke Sean M., Herskowitz Ira. A third osmosensing branch in Saccharomyces cerevisiae requires the Msb2 protein and functions in parallel with the Sho1 branch. Mol Cell Biol. 2002 Jul;22(13):4739–4749. doi: 10.1128/MCB.22.13.4739-4749.2002. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. O'Rourke Sean M., Herskowitz Ira, O'Shea Erin K. Yeast go the whole HOG for the hyperosmotic response. Trends Genet. 2002 Aug;18(8):405–412. doi: 10.1016/s0168-9525(02)02723-3. [DOI] [PubMed] [Google Scholar]
  30. Peter M., Gartner A., Horecka J., Ammerer G., Herskowitz I. FAR1 links the signal transduction pathway to the cell cycle machinery in yeast. Cell. 1993 May 21;73(4):747–760. doi: 10.1016/0092-8674(93)90254-n. [DOI] [PubMed] [Google Scholar]
  31. Philips J., Herskowitz I. Osmotic balance regulates cell fusion during mating in Saccharomyces cerevisiae. J Cell Biol. 1997 Sep 8;138(5):961–974. doi: 10.1083/jcb.138.5.961. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Phizicky E. M., Fields S. Protein-protein interactions: methods for detection and analysis. Microbiol Rev. 1995 Mar;59(1):94–123. doi: 10.1128/mr.59.1.94-123.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Posas F., Saito H. Osmotic activation of the HOG MAPK pathway via Ste11p MAPKKK: scaffold role of Pbs2p MAPKK. Science. 1997 Jun 13;276(5319):1702–1705. doi: 10.1126/science.276.5319.1702. [DOI] [PubMed] [Google Scholar]
  34. Printen J. A., Sprague G. F., Jr Protein-protein interactions in the yeast pheromone response pathway: Ste5p interacts with all members of the MAP kinase cascade. Genetics. 1994 Nov;138(3):609–619. doi: 10.1093/genetics/138.3.609. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Raitt D. C., Posas F., Saito H. Yeast Cdc42 GTPase and Ste20 PAK-like kinase regulate Sho1-dependent activation of the Hog1 MAPK pathway. EMBO J. 2000 Sep 1;19(17):4623–4631. doi: 10.1093/emboj/19.17.4623. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Roberts C. J., Nelson B., Marton M. J., Stoughton R., Meyer M. R., Bennett H. A., He Y. D., Dai H., Walker W. L., Hughes T. R. Signaling and circuitry of multiple MAPK pathways revealed by a matrix of global gene expression profiles. Science. 2000 Feb 4;287(5454):873–880. doi: 10.1126/science.287.5454.873. [DOI] [PubMed] [Google Scholar]
  37. Santos B., Duran A., Valdivieso M. H. CHS5, a gene involved in chitin synthesis and mating in Saccharomyces cerevisiae. Mol Cell Biol. 1997 May;17(5):2485–2496. doi: 10.1128/mcb.17.5.2485. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Santos B., Snyder M. Targeting of chitin synthase 3 to polarized growth sites in yeast requires Chs5p and Myo2p. J Cell Biol. 1997 Jan 13;136(1):95–110. doi: 10.1083/jcb.136.1.95. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Santos Beatriz, Snyder Michael. Specific protein targeting during cell differentiation: polarized localization of Fus1p during mating depends on Chs5p in Saccharomyces cerevisiae. Eukaryot Cell. 2003 Aug;2(4):821–825. doi: 10.1128/EC.2.4.821-825.2003. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Sheu Y. J., Santos B., Fortin N., Costigan C., Snyder M. Spa2p interacts with cell polarity proteins and signaling components involved in yeast cell morphogenesis. Mol Cell Biol. 1998 Jul;18(7):4053–4069. doi: 10.1128/mcb.18.7.4053. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Takei K., Slepnev V. I., Haucke V., De Camilli P. Functional partnership between amphiphysin and dynamin in clathrin-mediated endocytosis. Nat Cell Biol. 1999 May;1(1):33–39. doi: 10.1038/9004. [DOI] [PubMed] [Google Scholar]
  42. Tong Amy Hin Yan, Drees Becky, Nardelli Giuliano, Bader Gary D., Brannetti Barbara, Castagnoli Luisa, Evangelista Marie, Ferracuti Silvia, Nelson Bryce, Paoluzi Serena. A combined experimental and computational strategy to define protein interaction networks for peptide recognition modules. Science. 2001 Dec 13;295(5553):321–324. doi: 10.1126/science.1064987. [DOI] [PubMed] [Google Scholar]
  43. Trueheart J., Boeke J. D., Fink G. R. Two genes required for cell fusion during yeast conjugation: evidence for a pheromone-induced surface protein. Mol Cell Biol. 1987 Jul;7(7):2316–2328. doi: 10.1128/mcb.7.7.2316. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Trueheart J., Fink G. R. The yeast cell fusion protein FUS1 is O-glycosylated and spans the plasma membrane. Proc Natl Acad Sci U S A. 1989 Dec;86(24):9916–9920. doi: 10.1073/pnas.86.24.9916. [DOI] [PMC free article] [PubMed] [Google Scholar]
  45. Uetz P., Giot L., Cagney G., Mansfield T. A., Judson R. S., Knight J. R., Lockshon D., Narayan V., Srinivasan M., Pochart P. A comprehensive analysis of protein-protein interactions in Saccharomyces cerevisiae. Nature. 2000 Feb 10;403(6770):623–627. doi: 10.1038/35001009. [DOI] [PubMed] [Google Scholar]
  46. Zarzov P., Mazzoni C., Mann C. The SLT2(MPK1) MAP kinase is activated during periods of polarized cell growth in yeast. EMBO J. 1996 Jan 2;15(1):83–91. [PMC free article] [PubMed] [Google Scholar]
  47. Ziman M., Chuang J. S., Tsung M., Hamamoto S., Schekman R. Chs6p-dependent anterograde transport of Chs3p from the chitosome to the plasma membrane in Saccharomyces cerevisiae. Mol Biol Cell. 1998 Jun;9(6):1565–1576. doi: 10.1091/mbc.9.6.1565. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Genetics are provided here courtesy of Oxford University Press

RESOURCES