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Identification of protein kinase disruptions as suppressors of the calcium sensitivity of S. cerevisiae Δptp2 Δmsg5 protein phosphatase double disruptant

Archives of Microbiology volume 192, pages 157–165 (2010)Cite this article

Abstract

The double disruptant of the S. cerevisiae protein phosphatase (PPase) genes, PTP2 (phosphotyrosine-specific PPase) and MSG5 (phosphotyrosine and phosphothreonine/serine-PPase) causes calcium-sensitive growth (Cas). Previous study using Fluorescent-activated cell sorting (FACS) analysis showed that this growth defect with calcium occurs at G1–S transition in the cell cycle. We discovered that six non-essential protein kinase (PKase) disruptions (Δbck1, Δmkk1, Δslt2/Δmpk1, Δmck1, Δssk2 and Δyak1) suppressed the Cas-phenotype of the Δptp2 Δmsg5 double disruptant. Bck1p, Mkk1p and Slt2p are components of the mitogen-activated protein kinase (MAPK) cascade of cell wall integrity pathway (Slt2 pathway), and Mck1p is its down regulator. Ssk2p is the MAPK kinase kinase of the high-osmolarity glycerol (HOG) pathway, while Yak1p is a negative regulator for the cAMP-dependent PKA pathway. FACS analysis revealed that only the disruption of Δssk2 and Δyak1 but not Δbck1, Δmkk1, Δslt2 and Δmck1 was able to suppress the delayed G1–S transition, suggesting that suppression of the growth defect is not always accompanied by suppression of the G1–S transition delay. The discovery of these PKases as suppressors revealed that in addition to the previously anticipated Slt2 pathway, HOG, Yak1p and Mck1p regulatory pathways may also be involved in the calcium sensitivity of the Δptp2 Δmsg5 double disruptant.

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Abbreviations

PPase:

Protein phosphatase

PKase:

Protein kinase

MAPK:

Mitogen-activated protein kinase

Cas :

Calcium-sensitive growth

FACS:

Fluorescent-activated cell sorting

HOG:

High-osmolarity glycerol

References

  • Brachmann CB, Davies A, Cost GJ, Caputo E, Li J, Hieter P, Boeke JD (1998) Designer deletion strains derived from Saccharomyces cerevisiae S288C: a useful set of strains and plasmids for PCR-mediated gene disruption and other applications. Yeast 14:115–132

    Article  CAS  PubMed  Google Scholar 

  • Doi K, Gartner A, Ammerer G, Errede B, Shinkawa H, Sugimoto K, Matsumoto K (1994) MSG5, a novel protein phosphatase promotes adaptation to pheromone response in S.cerevisiae. EMBO J 13:61–70

    CAS  PubMed  Google Scholar 

  • Flandez M, Cosano IC, Nombela C, Martin H, Molina M (2004) Reciprocal regulation between Slt2 MAPK and isoforms of Msg5 dual-specificity protein phosphatase modulates the yeast cell integrity pathway. J Biol Chem 12:11027–11034

    Article  Google Scholar 

  • Guan KL, Deschenes RJ, Dixon JE (1992) Isolation and characterization of a second protein tyrosine phosphatase gene, PTP2, from Saccharomyces cerevisiae. J Biol Chem 267:10024–10030

    CAS  PubMed  Google Scholar 

  • Hermansyah H, Sugiyama M, Kaneko Y, Harashima S (2009) Yeast protein phosphatase Ptp2p and Msg5p are involved in G1-S transition, CLN2 transcription and vacuole morphogenesis. Arch Microbiol 191:721–733

    Article  CAS  PubMed  Google Scholar 

  • Hirasaki M, Kaneko Y, Harashima S (2008) Protein phosphatase Siw14 controls intracellular localization of Gln3 in cooperation with Npr1 kinase in Saccharomyces cerevisiae. Gene 409:34–43

    Article  CAS  PubMed  Google Scholar 

  • Hoffman-Sommer M, Migdalski A, Rytka J, Kucharczyk R (2005) Multiple functions of the vacuolar sorting protein Ccz1p in Saccharomyces cerevisiae. Biochem Biophys Res Commun 329:197–204

    Article  CAS  PubMed  Google Scholar 

  • Irie K, Takase M, Lee KS, Levin DE, Araki H, Matsumoto K, Oshima Y (1993) MKK1 and MKK2, which encode Saccharomyces cerevisiae mitogen-activated protein kinase-kinase homologs, function in the pathway mediated by protein kinase C. Mol Cell Biol 13:3076–3083

    CAS  PubMed  Google Scholar 

  • Jacoby T, Flanagan H, Faykin A, Seto AG, Mattison C, Ota I (1997) Two protein tyrosine phosphatase inactivate the osmotic stress response pathway in yeast by targeting the mitogen-activated protein kinase in Hog1. J Biol Chem 272:17749–17755

    Article  CAS  PubMed  Google Scholar 

  • Kitada K, Yamaguchi E, Arisawa M (1995) Cloning of the Candida glabrata TRP1 and HIS3 genes, and construction of their disruptant strains by sequential integrative transformation. Gene 165:203–206

    Article  CAS  PubMed  Google Scholar 

  • Lee P, Cho BR, Joo HS, Hahn JS (2008) Yeast Yak1 kinase, a bridge between PKA and stress-responsive transcription factors, Hsf1 and Msn2/Msn4. Mol Microbiol 70:882–895

    CAS  PubMed  Google Scholar 

  • Madden K, Sheu YJ, Baetz K, Andrews B, Snyder M (1997) SBF cell cycle regulator as a target of the yeast PKC-MAP kinase pathway. Science 275:1781–1784

    Article  CAS  PubMed  Google Scholar 

  • Martin H, Rodriguez-Pachon JM, Ruiz C, Nombela C, Molina M (2000) Regulatory mechanism for modulation of signaling through the cell integrity Slt2-mediated pathway in Saccharomyces cerevisiae. J Biol Chem 18:1511–1519

    Article  Google Scholar 

  • Mattison CP, Spencerm SS, Kresge KA, Lee J, Ota IM (1999) Differential regulation of the cell wall integrity mitogen-activated protein kinase pathway in the budding yeast by the protein tyrosine phosphatase Ptp2 and Ptp3. Mol Cell Biol 19:7651–7660

    CAS  PubMed  Google Scholar 

  • Mizunuma M, Hirata D, Miyahara K, Tsuchiya E, Miyakawa T (1998) Role of calcineurin and Mpk1 in regulating the onset of mitosis in budding yeast. Nature 392:303–306

    Article  CAS  PubMed  Google Scholar 

  • Mizunuma M, Hirata D, Miyaoka R, Miyakawa T (2001) GSK-3 kinase Mck1 and calcineurin coordinately mediate Hsl1 down-regulation by Ca2+ in budding yeast. EMBO J 20:1074–1085

    Article  CAS  PubMed  Google Scholar 

  • Mizunuma M, Hirata D, Miyakawa T (2005) Implication of Pkc1p protein kinase C in sustaining Cln2p level and polarized bud growth in response to calcium signaling in Saccharomyces cerevisiae. J Cell Sci 118:4219–4229

    Article  CAS  PubMed  Google Scholar 

  • Posas F, Saito H (1998) Activation of the yeast SSK2 MAP kinase kinase kinase by the SSK1 two component response regulator. EMBO J 17:1385–1394

    Article  CAS  PubMed  Google Scholar 

  • Rep M, Krantz M, Thevelein JM, Hohmann S (2000) The Hot1p and Msn2p/Msn4p are required for the induction of subsets of high osmolarity glycerol pathway-dependent genes. J Biol Chem 275:8290–8300

    Article  CAS  PubMed  Google Scholar 

  • Sakumoto N, Mukai Y, Uchida K, Kouchi T, Kuwajima J, Nakagawa Y, Sugioka S, Yamamoto E, Furuyama T, Mizubuchi H, Ohsugi N, Sakuno T, Kikuchi K, Matsuoka I, Ogawa N, Kaneko Y, Harashima S (1999) A series of protein phosphatase gene disruptants in Saccharomyces cerevisiae. Yeast 15:1669–1679

    Article  CAS  PubMed  Google Scholar 

  • Sakumoto N, Matsuoka I, Mukai Y, Ogawa N, Kaneko Y, Harashima Y (2002) A series of double disruptants for protein phosphatase genes in Saccharomyces cerevisiae and their phenotypic analysis. Yeast 19:587–599

    Article  CAS  PubMed  Google Scholar 

  • Seeley ES, Kato N, Margolis N, Wickner W, Eitzen G (2002) Genomic analysis of homotypic vacuole fusion. Mol Bio Cell 13:782–794

    Article  CAS  Google Scholar 

  • Sherman F, Hicks J (1991) Micromanipulation and dissection of Asci. In: Guide to yeast genetics and molecular biology. Methods Enzymol 194:21–37

  • Shitamukai A, Hirata D, Sonobe S, Miyakawa T (2004) Evidence for antagonistic regulation of cell growth by the calcineurin and high osmolarity glycerol pathways in Saccharomyces cerevisiae. J Biol Chem 279:3651–3661

    Article  CAS  PubMed  Google Scholar 

  • Smith A, Ward MP, Garrett S (1998) Yeast PKA represses Msn2p/Msn4p-dependent gene expression to regulate growth, stress response and glycogen accumulation. EMBO J 17:3556–3564

    Article  CAS  PubMed  Google Scholar 

  • Watanabe Y, Irie K, Matsumoto K (1995) Yeast RLM1 encodes a serum response factor-like protein that may function downstream of the Mpk1 (Slt2) mitogen-activated protein kinase pathway. Mol Cell Biol 15:5740–5749

    CAS  PubMed  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  CAS  PubMed  Google Scholar 

  • Winzeler EA, Shoemaker DD, Astromoff A, Liang H, Anderson K, Andre B, Bangham R, Benito R, Boeke JD, Bussey H, Chu AM, Connelly C, Davis K, Dietrich F, Dow SW, El Bakkoury M, Foury F, Friend SH, Gentalen E, Giaever G, Hegemann JH, Jones J, Laub M, Liao H, Liebundguth N, Lockhart DJ, Lucau-Danila A, Lussier M, M’Rabet N, Menard P, Mittmann M, Pai C, Rebischung C, Revuelta JL, Riles L, Roberts CJ, Ross-MacDonald P, Scherens B, Snyder M, Sookhai-Mahadeo S, Storms RK, Veronneau S, Voet M, Volckaert M, Ward TR, Wysocki R, Yen GS, Yu K, Zimmermann K, Philippsen P, Johnston M, Davis RW (1999) Functional characterization of the S. cerevisiae genome by gene deletion and parallel analysis. Science 285:901–906

    Article  CAS  PubMed  Google Scholar 

  • Zhan XL, Guan KL (1999) A specific protein-protein interaction accounts for the in vivo substrate selectivity of Ptp3 towards the Fus3 MAP kinase. Genes Dev 13:2811–2827

    Article  CAS  PubMed  Google Scholar 

  • Zhang J, Schneider C, Ottmers L, Rodriguez R, Day A, Markwardt J, Schneider BL (2002) Genomic scale mutant hunt identifies cell size homeostasis genes in S.cerevisiae. Curr Biol 12:1992–2001

    Article  CAS  PubMed  Google Scholar 

  • Zolnierowicz S, Bollen M (2000) Protein phosphorylation and protein phosphatases. EMBO J 19:483–488

    Article  CAS  PubMed  Google Scholar 

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Acknowledgments

This work was supported by a Grant-in-Aid for Scientific Research B, 2007–2009, to S.H. from the Ministry of Education, Science, Sports and Culture of Japan.

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Affiliations

  1. Department of Biotechnology, Graduate School of Engineering, Osaka University, 2-1 Yamadaoka, Suita, Osaka, 565-0871, Japan

    Hermansyah, Walter A. Laviña, Minetaka Sugiyama, Yoshinobu Kaneko & Satoshi Harashima

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  1. Hermansyah
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  2. Walter A. Laviña
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  3. Minetaka Sugiyama
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  4. Yoshinobu Kaneko
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  5. Satoshi Harashima
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Corresponding author

Correspondence to Satoshi Harashima.

Additional information

Communicated by Axel Brakhage.

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Hermansyah, Laviña, W.A., Sugiyama, M. et al. Identification of protein kinase disruptions as suppressors of the calcium sensitivity of S. cerevisiae Δptp2 Δmsg5 protein phosphatase double disruptant. Arch Microbiol 192, 157–165 (2010). https://doi.org/10.1007/s00203-009-0531-6

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  • DOI: https://doi.org/10.1007/s00203-009-0531-6

Keywords

  • PTP2
  • MSG5
  • Protein phosphatase
  • Protein Kinase
  • Calcium sensitive
  • Saccharomyces cerevisiae