Skip to main content
Log in

Regulation of respiratory growth by Ras: the glyoxylate cycle mutant, cit2Δ, is suppressed by RAS2

  • Research Article
  • Published:
Current Genetics Aims and scope Submit manuscript

Abstract

In Saccharomyces cerevisiae the Ras/cAMP/PKA signalling pathway controls multiple metabolic pathways, and alterations in the intracellular concentrations of cAMP through modification of signalling pathway factors can be lethal or result in severe growth defects. In this work, the important role of Ras2p in metabolic regulation during growth on the non-fermentable carbon source glycerol is further investigated. The data show that the overexpression of RAS2 suppresses the growth defect of the glyoxylate cycle citrate synthase mutant, cit2Δ. The overexpression results in enhanced proliferation and biomass yield when cells are grown on glycerol as sole carbon source, and increases citrate synthase activity and intracellular citrate concentration. Interestingly, the suppression of cit2Δ and the enhanced proliferation and biomass yield are only observed when RAS2 is overexpressed and not in strains containing the constitutively active allele RAS2 val19. However, both RAS2 and RAS2 val19upregulated citrate synthase activity. We propose that the RAS2 overexpression results in a combination of general upregulation of respiratory growth capacity and an increase in mitochondrial citrate/citrate synthases, which together, complement the metabolic requirements of the cit2Δ mutant. The data therefore provide new evidence for the role of Ras2p as a powerful modulator of metabolism during growth on a non-fermentable carbon source.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5

Similar content being viewed by others

References

  • Altschuler DL, Muro A, Schijman A, Almonacid FB, Torres HN (1990) Neurospora crassa cDNA clones coding for a new member of the ras protein family. FEBS Lett 273:103–106

    Article  PubMed  CAS  Google Scholar 

  • Becker DM, Guarente L (1991) High efficiency transformation of yeast by electroporation. In: Guthrie C, Fink GR (eds) Guide to yeast genetics and molecular biology. Academic, San Diego, pp 182–187

    Chapter  Google Scholar 

  • Bremer J (1983) Carnitine-metabolism and functions. Physiol Rev 63:1420–1480

    PubMed  CAS  Google Scholar 

  • Broach JR, Deschenes RJ (1990) The function of ras genes in Saccharomyces cerevisiae. Adv Cancer Res 54:79–139

    Article  PubMed  CAS  Google Scholar 

  • Brunelli JP, Pall ML (1993) A series of yeast/Escherichia coli lambda expression vectors designed for directional cloning of cDNAs and cre/lox-mediated plasmid excision. Yeast 9:1309–1318

    Article  PubMed  CAS  Google Scholar 

  • Chevtzoff C, Vallortigara J, Averet N, Rigoulet M, Devin A (2005) The yeast cAMP protein kinase Tpk3p is involved in the regulation of mitochondrial enzymatic content during growth. Biochim Biophys Acta 1706:117–125

    Article  PubMed  CAS  Google Scholar 

  • Colombo S, Ma P, Cauwenberg L, Winderickx J, Crauwels M, Teunissen A, Nauwelaers D, De Winde JH, Gorwa MF, Colavizza D, Thevelein JM (1998) Involvement of distinct G-proteins, Gpa2 and Ras, in glucose- and intracellular acidification-induced cAMP signalling in the yeast Saccharomyces cerevisiae. EMBO J 17:3326–3341

    Article  PubMed  CAS  Google Scholar 

  • De Vendittis E, Vitelli A, Zahn R, Fasano O (1986) Suppression of defective RAS1 and RAS2 functions in yeast by an adenylate cyclase activated by a single amino acid change. EMBO J 5:3657–3663

    PubMed  Google Scholar 

  • DeFeo-Jones D, Scolnick EM, Koller R, Dhar R (1983) ras-Related gene sequences identified and isolated from Saccharomyces cerevisiae. Nature 306:707–709

    Article  PubMed  CAS  Google Scholar 

  • Dejean L, Beauvoit B, Bunoust O, Guerin B, Rigoulet M (2002) Activation of Ras cascade increases the mitochondrial enzyme content of respiratory competent yeast. Biochem Biophys Res Commun 293:1383–1388

    Article  PubMed  CAS  Google Scholar 

  • Dhar R, Nieto A, Koller R, DeFeo-Jones D, Scolnick EM (1984) Nucleotide sequence of two rasH related-genes isolated from the yeast Saccharomyces cerevisiae. Nucleic Acids Res 12:3611–3618

    Article  PubMed  CAS  Google Scholar 

  • Engelberg D, Zandi E, Parker CS, Karin M (1994) The yeast and mammalian Ras pathways control transcription of heat shock genes independently of heat shock transcription factor. Mol Cell Biol 14:4929–4937

    PubMed  CAS  Google Scholar 

  • Field J, Nikawa J, Broek D, MacDonald B, Rodgers L, Wilson IA, Lerner RA, Wigler M (1988) Purification of a RAS-responsive adenylyl cyclase complex from Saccharomyces cerevisiae by use of an epitope addition method. Mol Cell Biol 8:2159–2165

    PubMed  CAS  Google Scholar 

  • Fraenkel DG (1985) On ras gene function in yeast. Proc Natl Acad Sci USA 82:4740–4744

    Article  PubMed  CAS  Google Scholar 

  • Hlavata L, Aguilaniu H, Pichova A, Nystrom T (2003) The oncogenic RAS2 val19 mutation locks respiration, independently of PKA, in a mode prone to generate ROS. EMBO J 22:3337–3345

    Article  PubMed  CAS  Google Scholar 

  • Jia YK, Becam AM, Herbert CJ (1997) The CIT3 gene of Saccharomyces cerevisiae encodes a second mitochondrial isoform of citrate synthase. Mol Microbiol 24:53–59

    Article  PubMed  CAS  Google Scholar 

  • Jones S, Vignais ML, Broach JR (1991) The CDC25 protein of Saccharomyces cerevisiae promotes exchange of guanine nucleotides bound to ras. Mol Cell Biol 11:2641–2646

    PubMed  CAS  Google Scholar 

  • Kataoka T, Powers S, McGill C, Fasano O, Strathern J, Broach J, Wigler M (1984) Genetic analysis of yeast RAS1 and RAS2 genes. Cell 37:437–445

    Article  PubMed  CAS  Google Scholar 

  • Kataoka T, Powers S, Cameron S, Fasano O, Goldfarb M, Broach J, Wigler M (1985) Functional homology of mammalian and yeast RAS genes. Cell 40:19–26

    Article  PubMed  CAS  Google Scholar 

  • Kim KS, Rosenkrantz MS, Guarente L (1986) Saccharomyces cerevisiae contains two functional citrate synthase genes. Mol Cell Biol 6:1936–1942

    PubMed  CAS  Google Scholar 

  • Kirchman PA, Kim S, Lai CY, Jazwinski SM (1999) Interorganelle signaling is a determinant of longevity in Saccharomyces cerevisiae. Genetics 152:179–190

    PubMed  CAS  Google Scholar 

  • Kornberg HL (1966) The role and control of the glyoxylate cycle in Escherichia coli. Biochem J 99:1–11

    PubMed  CAS  Google Scholar 

  • Lee BN, Elion EA (1999) The MAPKKK Ste11 regulates vegetative growth through a kinase cascade of shared signaling components. Proc Natl Acad Sci USA 96:12679–12684

    Article  PubMed  CAS  Google Scholar 

  • Liao XS, Small WC, Srere PA, Butow RA (1991) Intramitochondrial functions regulate nonmitochondrial citrate synthase (CIT2) expression in Saccharomyces cerevisiae. Mol Cell Biol 11:38–46

    PubMed  CAS  Google Scholar 

  • Mabuchi T, Ichimura Y, Takeda M, Douglas MG (2000) ASC1/RAS2 suppresses the growth defect on glycerol caused by the atp1–2 mutation in the yeast Saccharomyces cerevisiae. J Biol Chem 275:10492–10497

    Article  PubMed  CAS  Google Scholar 

  • Matsumoto K, Uno I, Oshima Y, Ishikawa T (1982) Isolation and characterization of yeast mutants deficient in adenylate cyclase and cAMP-dependent protein kinase. Proc Natl Acad Sci USA 79:2355–2359

    Article  PubMed  CAS  Google Scholar 

  • Mbonyi K, Van Aelst L, Arguelles JC, Jans AW, Thevelein JM (1990) Glucose-induced hyperaccumulation of cyclic AMP and defective glucose repression in yeast strains with reduced activity of cyclic AMP-dependent protein kinase. Mol Cell Biol 10:4518–4523

    PubMed  CAS  Google Scholar 

  • Mosch HU, Kubler E, Krappmann S, Fink GR, Braus GH (1999) Crosstalk between the Ras2p-controlled mitogen-activated protein kinase and cAMP pathways during invasive growth of Saccharomyces cerevisiae. Mol Biol Cell 10:1325–1335

    PubMed  CAS  Google Scholar 

  • Nikawa J, Cameron S, Toda T, Ferguson KM, Wigler M (1987) Rigorous feedback control of cAMP levels in Saccharomyces cerevisiae. Genes Dev 1:931–937

    Article  PubMed  CAS  Google Scholar 

  • Pan X, Heitman J (1999) Cyclic AMP-dependent protein kinase regulates pseudohyphal differentiation in Saccharomyces cerevisiae. Mol Cell Biol 19:4874–4887

    PubMed  CAS  Google Scholar 

  • Pan X, Harashima T, Heitman J (2000) Signal transduction cascades regulating pseudohyphal differentiation of Saccharomyces cerevisiae. Curr Opin Microbiol 3:567–572

    Article  PubMed  CAS  Google Scholar 

  • Parrini MC, Bernardi A, Parmeggiani A (1996) Determinants of Ras proteins specifying the sensitivity to yeast Ira2p and human p120-GAP. EMBO J 15:1107–1111

    PubMed  CAS  Google Scholar 

  • Pichova A, Vondrakova D, Breitenbach M (1997) Mutants in the Saccharomyces cerevisiae RAS2 gene influence life span, cytoskeleton, and regulation of mitosis. Can J Microbiol 43:774–781

    Article  PubMed  CAS  Google Scholar 

  • Powers S, Kataoka T, Fasano O, Goldfarb M, Strathern J, Broach J, Wigler M (1984) Genes in S. cerevisiae encoding proteins with domains homologous to the mammalian ras proteins. Cell 36:607–612

    Article  PubMed  CAS  Google Scholar 

  • Robertson LS, Causton HC, Young RA, Fink GR (2000) The yeast A kinases differentially regulate iron uptake and respiratory function. Proc Natl Acad Sci USA 97:5984–5988

    Article  PubMed  CAS  Google Scholar 

  • Rosenkrantz M, Alam T, Kim KS, Clark BJ, Srere PA, Guarente LP (1986) Mitochondrial and nonmitochondrial citrate synthases in Saccharomyces cerevisiae are encoded by distinct homologous genes. Mol Cell Biol 6:4509–4515

    PubMed  CAS  Google Scholar 

  • Russell M, Bradshaw-Rouse J, Markwardt D, Heideman W (1993) Changes in gene expression in the Ras/adenylate cyclase system of Saccharomyces cerevisiae: correlation with cAMP levels and growth arrest. Mol Biol Cell 4:757–765

    PubMed  CAS  Google Scholar 

  • Sass P, Field J, Nikawa J, Toda T, Wigler M (1986) Cloning and characterization of the high-affinity cAMP phosphodiesterase of Saccharomyces cerevisiae. Proc Natl Acad Sci USA 83:9303–9307

    Article  PubMed  CAS  Google Scholar 

  • Shama S, Kirchman PA, Jiang JC, Jazwinski SM (1998) Role of RAS2 in recovery from chronic stress: effect on yeast life span. Exp Cell Res 245:368–378

    Article  PubMed  CAS  Google Scholar 

  • Srere PA (1969) Citrate synthase. Meth Enzymol 13:3–11

    Article  CAS  Google Scholar 

  • Stanhill A, Schick N, Engelberg D (1999) The yeast ras/cyclic AMP pathway induces invasive growth by suppressing the cellular stress response. Mol Cell Biol 19:7529–7538

    PubMed  CAS  Google Scholar 

  • Steinmetz LM, Scharfe C, Deutschbauer AM, Mokranjac D, Herman ZS, Jones T, Chu AM, Giaever G, Prokisch H, Oefner PJ, Davis RW (2002) Systematic screen for human disease genes in yeast. Nat Genet 31:400–404

    PubMed  CAS  Google Scholar 

  • Sun J, Kale SP, Childress AM, Pinswasdi C, Jazwinski SM (1994) Divergent roles of RAS1 and RAS2 in yeast longevity. J Biol Chem 269:18638–18645

    PubMed  CAS  Google Scholar 

  • Swiegers JH, Dippenaar N, Pretorius IS, Bauer FF (2001) Carnitine-dependent metabolic activities in Saccharomyces cerevisiae: three carnitine acetyltransferases are essential in a carnitine-dependent strain. Yeast 18:585–595

    Article  PubMed  CAS  Google Scholar 

  • Tanaka K, Matsumoto K, Toh-E A (1989) IRA1, an inhibitory regulator of the RAS-cyclic AMP pathway in Saccharomyces cerevisiae. Mol Cell Biol 9:757–768

    PubMed  CAS  Google Scholar 

  • Tanaka K, Nakafuku M, Satoh T, Marshall MS, Gibbs JB, Matsumoto K, Kaziro Y, Toh-e A (1990a) S. cerevisiae genes IRA1 and IRA2 encode proteins that may be functionally equivalent to mammalian ras GTPase activating protein. Cell 60:803–807

    Article  CAS  Google Scholar 

  • Tanaka K, Nakafuku M, Tamanoi F, Kaziro Y, Matsumoto K, Toh-e A (1990b) IRA2, a second gene of Saccharomyces cerevisiae that encodes a protein with a domain homologous to mammalian ras GTPase-activating protein. Mol Cell Biol 10:4303–4313

    CAS  Google Scholar 

  • Tatchell K, Chaleff DT, DeFeo-Jones D, Scolnick EM (1984) Requirement of either of a pair of ras-related genes of Saccharomyces cerevisiae for spore viability. Nature 309:523–527

    Article  PubMed  CAS  Google Scholar 

  • Tatchell K, Robinson LC, Breitenbach M (1985) RAS2 of Saccharomyces cerevisiae is required for gluconeogenic growth and proper response to nutrient limitation. Proc Natl Acad Sci USA 82:3785–3789

    Article  PubMed  CAS  Google Scholar 

  • Thevelein JM (1994) Signal transduction in yeast. Yeast 10:1753–1790

    Article  PubMed  CAS  Google Scholar 

  • Toda T, Uno I, Ishikawa T, Powers S, Kataoka T, Broek D, Cameron S, Broach J, Matsumoto K, Wigler M (1985) In yeast, RAS proteins are controlling elements of adenylate cyclase. Cell 40:27–36

    Article  PubMed  CAS  Google Scholar 

  • Toda T, Cameron S, Sass P, Zoller M, Scott JD, McMullen B, Hurwitz M, Krebs EG, Wigler M (1987a) Cloning and characterization of BCY1, a locus encoding a regulatory subunit of the cyclic AMP-dependent protein kinase in Saccharomyces cerevisiae. Mol Cell Biol 7:1371–1377

    CAS  Google Scholar 

  • Toda T, Cameron S, Sass P, Zoller M, Wigler M (1987b) Three different genes in S. cerevisiae encode the catalytic subunits of the cAMP-dependent protein kinase. Cell 50:277–287

    Article  CAS  Google Scholar 

  • Van Roermund CW, Elgersma Y, Singh N, Wanders RJ, Tabak HF (1995) The membrane of peroxisomes in Saccharomyces cerevisiae is impermeable to NAD(H) and acetyl-CoA under in vivo conditions. EMBO J 14:3480–3486

    PubMed  Google Scholar 

  • Volschenk H, Viljoen M, Grobler J, Petzold B, Bauer F, Subden RE, Young RA, Lonvaud A, Denayrolles M, Van Vuuren HJ (1997) Engineering pathways for malate degradation in Saccharomyces cerevisiae. Nat Biotechnol 15:253–257

    Article  PubMed  CAS  Google Scholar 

  • Winston F, Dollard C, Ricupero-Hovasse SL (1995) Construction of a set of convenient Saccharomyces cerevisiae strains that are isogenic to S288C. Yeast 11:53–55

    Article  PubMed  CAS  Google Scholar 

Download references

Acknowledgements

We would like to thank the South African National Research Foundation and Winetech for financial support. We are grateful to David Engelberg for supplying the B2562 plasmid.

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to Jan H. Swiegers.

Additional information

Communicated by S. Hohmann

Rights and permissions

Reprints and permissions

About this article

Cite this article

Swiegers, J.H., Pretorius, I.S. & Bauer, F.F. Regulation of respiratory growth by Ras: the glyoxylate cycle mutant, cit2Δ, is suppressed by RAS2 . Curr Genet 50, 161–171 (2006). https://doi.org/10.1007/s00294-006-0084-z

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00294-006-0084-z

Keywords

Navigation