Skip to main content
PMC home page
The EMBO Journal logoLink to The EMBO Journal
. 1997 Dec 1;16(23):7008–7018. doi: 10.1093/emboj/16.23.7008

Yeast pseudohyphal growth is regulated by GPA2, a G protein alpha homolog.

M C Lorenz 1, J Heitman 1
PMCID: PMC1170304  PMID: 9384580

Abstract

Pseudohyphal differentiation, a filamentous growth form of the budding yeast Saccharomyces cerevisiae, is induced by nitrogen starvation. The mechanisms by which nitrogen limitation regulates this process are currently unknown. We have found that GPA2, one of the two heterotrimeric G protein alpha subunit homologs in yeast, regulates pseudohyphal differentiation. Deltagpa2/Deltagpa2 mutant strains have a defect in pseudohyphal growth. In contrast, a constitutively active allele of GPA2 stimulates filamentation, even on nitrogen-rich media. Moreover, a dominant negative GPA2 allele inhibits filamentation of wild-type strains. Several findings, including epistasis analysis and reporter gene studies, indicate that GPA2 does not regulate the MAP kinase cascade known to regulate filamentous growth. Previous studies have implicated GPA2 in the control of intracellular cAMP levels; we find that expression of the dominant RAS2(Gly19Val) mutant or exogenous cAMP suppresses the Deltagpa2 pseudohyphal defect. cAMP also stimulates filamentation in strains lacking the cAMP phosphodiesterase PDE2, even in the absence of nitrogen starvation. Our findings suggest that GPA2 is an element of the nitrogen sensing machinery that regulates pseudohyphal differentiation by modulating cAMP levels.

Full Text

The Full Text of this article is available as a PDF (504.1 KB).

Selected References

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

  1. Akada R., Kallal L., Johnson D. I., Kurjan J. Genetic relationships between the G protein beta gamma complex, Ste5p, Ste20p and Cdc42p: investigation of effector roles in the yeast pheromone response pathway. Genetics. 1996 May;143(1):103–117. doi: 10.1093/genetics/143.1.103. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Alani E., Cao L., Kleckner N. A method for gene disruption that allows repeated use of URA3 selection in the construction of multiply disrupted yeast strains. Genetics. 1987 Aug;116(4):541–545. doi: 10.1534/genetics.112.541.test. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Arkinstall S. J., Papasavvas S. G., Payton M. A. Yeast alpha-mating factor receptor-linked G-protein signal transduction suppresses Ras-dependent activity. FEBS Lett. 1991 Jun 17;284(1):123–128. doi: 10.1016/0014-5793(91)80777-z. [DOI] [PubMed] [Google Scholar]
  4. Bardwell L., Cook J. G., Inouye C. J., Thorner J. Signal propagation and regulation in the mating pheromone response pathway of the yeast Saccharomyces cerevisiae. Dev Biol. 1994 Dec;166(2):363–379. doi: 10.1006/dbio.1994.1323. [DOI] [PubMed] [Google Scholar]
  5. Broach J. R., Deschenes R. J. The function of ras genes in Saccharomyces cerevisiae. Adv Cancer Res. 1990;54:79–139. doi: 10.1016/s0065-230x(08)60809-x. [DOI] [PubMed] [Google Scholar]
  6. Field J., Nikawa J., Broek D., MacDonald B., Rodgers L., Wilson I. A., Lerner R. A., Wigler M. Purification of a RAS-responsive adenylyl cyclase complex from Saccharomyces cerevisiae by use of an epitope addition method. Mol Cell Biol. 1988 May;8(5):2159–2165. doi: 10.1128/mcb.8.5.2159. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Gavrias V., Andrianopoulos A., Gimeno C. J., Timberlake W. E. Saccharomyces cerevisiae TEC1 is required for pseudohyphal growth. Mol Microbiol. 1996 Mar;19(6):1255–1263. doi: 10.1111/j.1365-2958.1996.tb02470.x. [DOI] [PubMed] [Google Scholar]
  8. Gietz R. D., Sugino A. New yeast-Escherichia coli shuttle vectors constructed with in vitro mutagenized yeast genes lacking six-base pair restriction sites. Gene. 1988 Dec 30;74(2):527–534. doi: 10.1016/0378-1119(88)90185-0. [DOI] [PubMed] [Google Scholar]
  9. Gimeno C. J., Fink G. R. Induction of pseudohyphal growth by overexpression of PHD1, a Saccharomyces cerevisiae gene related to transcriptional regulators of fungal development. Mol Cell Biol. 1994 Mar;14(3):2100–2112. doi: 10.1128/mcb.14.3.2100. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Gimeno C. J., Ljungdahl P. O., Styles C. A., Fink G. R. Unipolar cell divisions in the yeast S. cerevisiae lead to filamentous growth: regulation by starvation and RAS. Cell. 1992 Mar 20;68(6):1077–1090. doi: 10.1016/0092-8674(92)90079-r. [DOI] [PubMed] [Google Scholar]
  11. Gold S., Duncan G., Barrett K., Kronstad J. cAMP regulates morphogenesis in the fungal pathogen Ustilago maydis. Genes Dev. 1994 Dec 1;8(23):2805–2816. doi: 10.1101/gad.8.23.2805. [DOI] [PubMed] [Google Scholar]
  12. Graziano M. P., Gilman A. G. Synthesis in Escherichia coli of GTPase-deficient mutants of Gs alpha. J Biol Chem. 1989 Sep 15;264(26):15475–15482. [PubMed] [Google Scholar]
  13. Grenson M., Mousset M., Wiame J. M., Bechet J. Multiplicity of the amino acid permeases in Saccharomyces cerevisiae. I. Evidence for a specific arginine-transporting system. Biochim Biophys Acta. 1966 Oct 31;127(2):325–338. doi: 10.1016/0304-4165(66)90387-4. [DOI] [PubMed] [Google Scholar]
  14. Hartmann H. A., Kahmann R., Bölker M. The pheromone response factor coordinates filamentous growth and pathogenicity in Ustilago maydis. EMBO J. 1996 Apr 1;15(7):1632–1641. [PMC free article] [PubMed] [Google Scholar]
  15. Ho S. N., Hunt H. D., Horton R. M., Pullen J. K., Pease L. R. Site-directed mutagenesis by overlap extension using the polymerase chain reaction. Gene. 1989 Apr 15;77(1):51–59. doi: 10.1016/0378-1119(89)90358-2. [DOI] [PubMed] [Google Scholar]
  16. Isshiki T., Mochizuki N., Maeda T., Yamamoto M. Characterization of a fission yeast gene, gpa2, that encodes a G alpha subunit involved in the monitoring of nutrition. Genes Dev. 1992 Dec;6(12B):2455–2462. doi: 10.1101/gad.6.12b.2455. [DOI] [PubMed] [Google Scholar]
  17. Ivey F. D., Hodge P. N., Turner G. E., Borkovich K. A. The G alpha i homologue gna-1 controls multiple differentiation pathways in Neurospora crassa. Mol Biol Cell. 1996 Aug;7(8):1283–1297. doi: 10.1091/mbc.7.8.1283. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Kim D. U., Park S. K., Chung K. S., Choi M. U., Yoo H. S. The G protein beta subunit Gpb1 of Schizosaccharomyces pombe is a negative regulator of sexual development. Mol Gen Genet. 1996 Aug 27;252(1-2):20–32. doi: 10.1007/BF02173201. [DOI] [PubMed] [Google Scholar]
  19. Kozak K. R., Foster L. M., Ross I. K. Cloning and characterization of a G protein alpha-subunit-encoding gene from the basidiomycete, Coprinus congregatus. Gene. 1995 Sep 22;163(1):133–137. doi: 10.1016/0378-1119(95)00378-j. [DOI] [PubMed] [Google Scholar]
  20. Kron S. J., Styles C. A., Fink G. R. Symmetric cell division in pseudohyphae of the yeast Saccharomyces cerevisiae. Mol Biol Cell. 1994 Sep;5(9):1003–1022. doi: 10.1091/mbc.5.9.1003. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Kurjan J., Hirsch J. P., Dietzel C. Mutations in the guanine nucleotide-binding domains of a yeast G alpha protein confer a constitutive or uninducible state to the pheromone response pathway. Genes Dev. 1991 Mar;5(3):475–483. doi: 10.1101/gad.5.3.475. [DOI] [PubMed] [Google Scholar]
  22. Kurjan J. The pheromone response pathway in Saccharomyces cerevisiae. Annu Rev Genet. 1993;27:147–179. doi: 10.1146/annurev.ge.27.120193.001051. [DOI] [PubMed] [Google Scholar]
  23. Köhler J. R., Fink G. R. Candida albicans strains heterozygous and homozygous for mutations in mitogen-activated protein kinase signaling components have defects in hyphal development. Proc Natl Acad Sci U S A. 1996 Nov 12;93(23):13223–13228. doi: 10.1073/pnas.93.23.13223. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Kübler E., Mösch H. U., Rupp S., Lisanti M. P. Gpa2p, a G-protein alpha-subunit, regulates growth and pseudohyphal development in Saccharomyces cerevisiae via a cAMP-dependent mechanism. J Biol Chem. 1997 Aug 15;272(33):20321–20323. doi: 10.1074/jbc.272.33.20321. [DOI] [PubMed] [Google Scholar]
  25. Laloux I., Jacobs E., Dubois E. Involvement of SRE element of Ty1 transposon in TEC1-dependent transcriptional activation. Nucleic Acids Res. 1994 Mar 25;22(6):999–1005. doi: 10.1093/nar/22.6.999. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Leberer E., Harcus D., Broadbent I. D., Clark K. L., Dignard D., Ziegelbauer K., Schmidt A., Gow N. A., Brown A. J., Thomas D. Y. Signal transduction through homologs of the Ste20p and Ste7p protein kinases can trigger hyphal formation in the pathogenic fungus Candida albicans. Proc Natl Acad Sci U S A. 1996 Nov 12;93(23):13217–13222. doi: 10.1073/pnas.93.23.13217. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Leeuw T., Fourest-Lieuvin A., Wu C., Chenevert J., Clark K., Whiteway M., Thomas D. Y., Leberer E. Pheromone response in yeast: association of Bem1p with proteins of the MAP kinase cascade and actin. Science. 1995 Nov 17;270(5239):1210–1213. doi: 10.1126/science.270.5239.1210. [DOI] [PubMed] [Google Scholar]
  28. Liang H., Gaber R. F. A novel signal transduction pathway in Saccharomyces cerevisiae defined by Snf3-regulated expression of HXT6. Mol Biol Cell. 1996 Dec;7(12):1953–1966. doi: 10.1091/mbc.7.12.1953. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Liu H., Köhler J., Fink G. R. Suppression of hyphal formation in Candida albicans by mutation of a STE12 homolog. Science. 1994 Dec 9;266(5191):1723–1726. doi: 10.1126/science.7992058. [DOI] [PubMed] [Google Scholar]
  30. Liu H., Styles C. A., Fink G. R. Elements of the yeast pheromone response pathway required for filamentous growth of diploids. Science. 1993 Dec 10;262(5140):1741–1744. doi: 10.1126/science.8259520. [DOI] [PubMed] [Google Scholar]
  31. Lowy D. R., Willumsen B. M. Function and regulation of ras. Annu Rev Biochem. 1993;62:851–891. doi: 10.1146/annurev.bi.62.070193.004223. [DOI] [PubMed] [Google Scholar]
  32. Madhani H. D., Fink G. R. Combinatorial control required for the specificity of yeast MAPK signaling. Science. 1997 Feb 28;275(5304):1314–1317. doi: 10.1126/science.275.5304.1314. [DOI] [PubMed] [Google Scholar]
  33. Maeda T., Mochizuki N., Yamamoto M. Adenylyl cyclase is dispensable for vegetative cell growth in the fission yeast Schizosaccharomyces pombe. Proc Natl Acad Sci U S A. 1990 Oct;87(20):7814–7818. doi: 10.1073/pnas.87.20.7814. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Masters S. B., Miller R. T., Chi M. H., Chang F. H., Beiderman B., Lopez N. G., Bourne H. R. Mutations in the GTP-binding site of GS alpha alter stimulation of adenylyl cyclase. J Biol Chem. 1989 Sep 15;264(26):15467–15474. [PubMed] [Google Scholar]
  35. Medoff G., Maresca B., Lambowitz A. M., Kobayashi G., Painter A., Sacco M., Carratu L. Correlation between pathogenicity and temperature sensitivity in different strains of Histoplasma capsulatum. J Clin Invest. 1986 Dec;78(6):1638–1647. doi: 10.1172/JCI112757. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Miller R. T., Masters S. B., Sullivan K. A., Beiderman B., Bourne H. R. A mutation that prevents GTP-dependent activation of the alpha chain of Gs. Nature. 1988 Aug 25;334(6184):712–715. doi: 10.1038/334712a0. [DOI] [PubMed] [Google Scholar]
  37. Miyajima I., Nakafuku M., Nakayama N., Brenner C., Miyajima A., Kaibuchi K., Arai K., Kaziro Y., Matsumoto K. GPA1, a haploid-specific essential gene, encodes a yeast homolog of mammalian G protein which may be involved in mating factor signal transduction. Cell. 1987 Sep 25;50(7):1011–1019. doi: 10.1016/0092-8674(87)90167-x. [DOI] [PubMed] [Google Scholar]
  38. Mösch H. U., Fink G. R. Dissection of filamentous growth by transposon mutagenesis in Saccharomyces cerevisiae. Genetics. 1997 Mar;145(3):671–684. doi: 10.1093/genetics/145.3.671. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Mösch H. U., Roberts R. L., Fink G. R. Ras2 signals via the Cdc42/Ste20/mitogen-activated protein kinase module to induce filamentous growth in Saccharomyces cerevisiae. Proc Natl Acad Sci U S A. 1996 May 28;93(11):5352–5356. doi: 10.1073/pnas.93.11.5352. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Nakafuku M., Obara T., Kaibuchi K., Miyajima I., Miyajima A., Itoh H., Nakamura S., Arai K., Matsumoto K., Kaziro Y. Isolation of a second yeast Saccharomyces cerevisiae gene (GPA2) coding for guanine nucleotide-binding regulatory protein: studies on its structure and possible functions. Proc Natl Acad Sci U S A. 1988 Mar;85(5):1374–1378. doi: 10.1073/pnas.85.5.1374. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Neer E. J. Heterotrimeric G proteins: organizers of transmembrane signals. Cell. 1995 Jan 27;80(2):249–257. doi: 10.1016/0092-8674(95)90407-7. [DOI] [PubMed] [Google Scholar]
  42. Niimi M., Niimi K., Tokunaga J., Nakayama H. Changes in cyclic nucleotide levels and dimorphic transition in Candida albicans. J Bacteriol. 1980 Jun;142(3):1010–1014. doi: 10.1128/jb.142.3.1010-1014.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Obara T., Nakafuku M., Yamamoto M., Kaziro Y. Isolation and characterization of a gene encoding a G-protein alpha subunit from Schizosaccharomyces pombe: involvement in mating and sporulation pathways. Proc Natl Acad Sci U S A. 1991 Jul 1;88(13):5877–5881. doi: 10.1073/pnas.88.13.5877. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Ozcan S., Dover J., Rosenwald A. G., Wölfl S., Johnston M. Two glucose transporters in Saccharomyces cerevisiae are glucose sensors that generate a signal for induction of gene expression. Proc Natl Acad Sci U S A. 1996 Oct 29;93(22):12428–12432. doi: 10.1073/pnas.93.22.12428. [DOI] [PMC free article] [PubMed] [Google Scholar]
  45. Papasavvas S., Arkinstall S., Reid J., Payton M. Yeast alpha-mating factor receptor and G-protein-linked adenylyl cyclase inhibition requires RAS2 and GPA2 activities. Biochem Biophys Res Commun. 1992 May 15;184(3):1378–1385. doi: 10.1016/s0006-291x(05)80035-x. [DOI] [PubMed] [Google Scholar]
  46. Paznokas J. L., Sypherd P. S. Respiratory capacity, cyclic adenosine 3',5'-monophosphate, and morphogenesis of Mucor racemosus. J Bacteriol. 1975 Oct;124(1):134–139. doi: 10.1128/jb.124.1.134-139.1975. [DOI] [PMC free article] [PubMed] [Google Scholar]
  47. Regenfelder E., Spellig T., Hartmann A., Lauenstein S., Bölker M., Kahmann R. G proteins in Ustilago maydis: transmission of multiple signals? EMBO J. 1997 Apr 15;16(8):1934–1942. doi: 10.1093/emboj/16.8.1934. [DOI] [PMC free article] [PubMed] [Google Scholar]
  48. Sabie F. T., Gadd G. M. Effect of nucleosides and nucleotides and the relationship between cellular adenosine 3':5'-cyclic monophosphate (cyclic AMP) and germ tube formation in Candida albicans. Mycopathologia. 1992 Sep;119(3):147–156. doi: 10.1007/BF00448812. [DOI] [PubMed] [Google Scholar]
  49. Sherman F. Getting started with yeast. Methods Enzymol. 1991;194:3–21. doi: 10.1016/0076-6879(91)94004-v. [DOI] [PubMed] [Google Scholar]
  50. Stevenson B. J., Rhodes N., Errede B., Sprague G. F., Jr Constitutive mutants of the protein kinase STE11 activate the yeast pheromone response pathway in the absence of the G protein. Genes Dev. 1992 Jul;6(7):1293–1304. doi: 10.1101/gad.6.7.1293. [DOI] [PubMed] [Google Scholar]
  51. Toda T., Uno I., Ishikawa T., Powers S., Kataoka T., Broek D., Cameron S., Broach J., Matsumoto K., Wigler M. In yeast, RAS proteins are controlling elements of adenylate cyclase. Cell. 1985 Jan;40(1):27–36. doi: 10.1016/0092-8674(85)90305-8. [DOI] [PubMed] [Google Scholar]
  52. Wach A., Brachat A., Pöhlmann R., Philippsen P. New heterologous modules for classical or PCR-based gene disruptions in Saccharomyces cerevisiae. Yeast. 1994 Dec;10(13):1793–1808. doi: 10.1002/yea.320101310. [DOI] [PubMed] [Google Scholar]
  53. Ward M. P., Gimeno C. J., Fink G. R., Garrett S. SOK2 may regulate cyclic AMP-dependent protein kinase-stimulated growth and pseudohyphal development by repressing transcription. Mol Cell Biol. 1995 Dec;15(12):6854–6863. doi: 10.1128/mcb.15.12.6854. [DOI] [PMC free article] [PubMed] [Google Scholar]
  54. Whiteway M., Hougan L., Dignard D., Thomas D. Y., Bell L., Saari G. C., Grant F. J., O'Hara P., MacKay V. L. The STE4 and STE18 genes of yeast encode potential beta and gamma subunits of the mating factor receptor-coupled G protein. Cell. 1989 Feb 10;56(3):467–477. doi: 10.1016/0092-8674(89)90249-3. [DOI] [PubMed] [Google Scholar]
  55. Yarden O., Plamann M., Ebbole D. J., Yanofsky C. cot-1, a gene required for hyphal elongation in Neurospora crassa, encodes a protein kinase. EMBO J. 1992 Jun;11(6):2159–2166. doi: 10.1002/j.1460-2075.1992.tb05275.x. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from The EMBO Journal are provided here courtesy of Nature Publishing Group

RESOURCES