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Protective roles and Pap1-dependent regulation of the Schizosaccharomyces pombe spy1 gene under nitrosative and nutritional stresses

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Abstract

This work aimed to assess novel protective roles and regulation of Spy1, a histidine-containing phosphotransfer (HPt) protein, in the fission yeast Schizosaccharomyces pombe. The structural gene encoding Spy1 was cloned into the shuttle vector pRS316 to generate the recombinant plasmid pYFSpy1. The spy1 + mRNA level was notably increased in S. pombe cells harboring the plasmid pYFSpy1. The S. pombe cells harboring pYFSpy1 exhibited higher survival than the vector control cells on the minimal media plates with nitric oxide (NO)-generating sodium nitroprusside (SNP) or without nitrogen source. In the liquid minimal media, they also showed higher viability under nitrosative stress or nitrogen-starved condition. The intracellular reactive oxygen species (ROS) level appeared to be lower in the fission yeast cells harboring pYFSpy1 than in the control yeast cells. Overexpression of the spy1 + gene showed scavenging effect on NO generated from SNP. Synthesis of β-galactosidase from the spy1 +-lacZ fusion gene was significantly enhanced by SNP and nitrogen starvation in the Pap1-positive but not in the Pap1-negative cells. The spy1 + mRNA level in S. pombe was also elevated by SNP and nitrogen starvation in the Pap1-positive but not in the Pap1-negative cells. In summary, Spy1 plays protective roles against nitrosative and nutritional stress in the fission yeast and is transcriptionally up-regulated by nitrosative and nutritional stresses in a Pap1-dependent manner.

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Abbreviations

NO:

Nitric oxide

ONPG:

ο-Nitrophenyl-β-d-galactopyranoside

PCR:

Polymerase chain reaction

ROS:

Reactive oxygen species

RT-PCR:

Reverse transcriptase-polymerase chain reaction

SNP:

Sodium nitroprusside

S. pombe :

Schizosaccharomyces pombe

References

  1. Imamura A, Hanaki N, Umeda H, Nakamura A, Suzuki T, Ueguchi C, Mizuno T (1998) Response regulators implicated in His-to-Asp phosphotransfer signaling in Arabidopsis. Proc Natl Acad Sci USA 95:2691–2696

    Article  CAS  PubMed  Google Scholar 

  2. Lu JM, Deschenes RJ, Fassler JS (2003) Saccharomyces cerevisiae histidine phosphotransferase Ypd1p shuttles between the nucleus and cytoplasm for SLN1-dependent phosphorylation of Ssk1p and Skn7p. Eukaryot Cell 2:1304–1314

    Article  CAS  PubMed  Google Scholar 

  3. Porter SW, West AH (2005) A common docking site for response regulators on the yeast phosphorelay protein YPD1. Biochim Biophys Acta 1748:138–145

    CAS  PubMed  Google Scholar 

  4. Aoyama K, Mitsubayashi Y, Aiba H, Mizuno T (2000) Spy1, a histidine-containing phosphotransfer signaling protein, regulates the fission yeast cell cycle through the Mcs4 response regulator. J Bacteriol 182:4868–4874

    Article  CAS  PubMed  Google Scholar 

  5. Aoyama K, Aiba H, Mizuno T (2001) Genetic analysis of the His-to-Asp phosphorelay implicated in mitotic cell cycle control: involvement of histidine-kinase genes of Schizosaccharomyces pombe. Biosci Biotechnol Biochem 65:2347–2352

    Article  CAS  PubMed  Google Scholar 

  6. Takatsume Y, Izawa S, Inoue Y (2006) Methylglyoxal as a signal initiator for activation of the stress-activated protein kinase cascade in the fission yeast Schizosaccharomyces pombe. J Biol Chem 281:9086–9092

    Article  CAS  PubMed  Google Scholar 

  7. Lenormand JL, Dellinger RW, Knudsen KE, Subramani S, Donoghue DJ (1999) Speedy: a novel cell cycle regulator of the G2/M transition. EMBO J 18:1869–1877

    Article  CAS  PubMed  Google Scholar 

  8. Toone WM, Kuge S, Samuels M, Morgan BA, Toda T, Jones N (1998) Regulation of the fission yeast transcription factor Pap1 by oxidative stress: requirement for the nuclear export factor Crm1 (exportin) and the stress-activated MAP kinase Sty1/Spc1. Genes Dev 12:1453–1463

    Article  CAS  PubMed  Google Scholar 

  9. Fujii Y, Shimizu T, Toda T, Yanagida M, Hakoshima T (2000) Structural basis for the diversity of DNA recognition by bZIP transcription factors. Nat Struct Biol 7:889–893

    Article  CAS  PubMed  Google Scholar 

  10. Toda T, Shimanuki M, Yanagida M (1991) Fission yeast genes that confer resistance to staurosporine encodes an AP-1 like transcription factor and a protein kinase related to the mammalian ERK1/MAP2 and budding yeast FUS3 and KSS1 kinases. Gens Dev 5:60–73

    Article  CAS  Google Scholar 

  11. Kang MH, Park EH, Lim CJ (2007) Protective role and regulation of Rad9 from the fission yeast Schizosaccharomyces pombe. FEMS Microbiol Lett 275:270–277

    Article  CAS  PubMed  Google Scholar 

  12. Kang GY, Park EH, Lim CJ (2008) Molecular cloning, characterization and regulation of a peroxiredoxin gene from Schizosaccharomyces pombe. Mol Biol Rep 35:387–395

    Article  CAS  PubMed  Google Scholar 

  13. Lee EH, Hyun DH, Park EH, Lim CJ (2010) A second protein disulfide isomerase plays a protective role against nitrosative and nutritional stresses in Schizosaccharomyces pombe. Mol Biol Rep. doi:10.1007/s11033-010-0018-1

  14. Kim SJ, Choi YS, Kim HG, Park EH, Lim CJ (2006) Cloning, characterization and regulation of a protein disulfide isomerase from the fission yeast Schizosaccharomyces pombe. Mol Biol Rep 33:187–196

    Article  CAS  PubMed  Google Scholar 

  15. Kim HJ, Jung HY, Lim CJ (2008) The pap1+ gene of fission yeast is transcriptionally regulated by nitrosative and nutritional stresses. FEMS Microbiol Lett 280:176–181

    Article  CAS  PubMed  Google Scholar 

  16. Myers AM, Tzagoloff A, Kinney DM, Lusty CJ (1986) Yeast shuttle and integrative vectors with multiple cloning sites suitable for construction of lacZ fusions. Gene 45:299–310

    Article  CAS  PubMed  Google Scholar 

  17. Guarente L (1983) Yeast promoters and lacZ fusions designed to study expression of cloned genes in yeast. Methods Enzymol 101:181–191

    Article  CAS  PubMed  Google Scholar 

  18. Bradford MM (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 72:248–254

    Article  CAS  PubMed  Google Scholar 

  19. Sherman MP, Aeberhard EE, Wong VZ, Griscavage JM, Ignarro LJ (1993) Pyrrolidine dithiocarbamate inhibits induction of nitric oxide synthase activity in rat alveolar macrophages. Biochem Biophys Res Commun 191:1301–1308

    Article  CAS  PubMed  Google Scholar 

  20. Royall JA, Ischiropoulos H (1993) Evaluation of 2′,7′-dichlorofluorescin and dihydrorhodamine 123 as fluorescent probes for intracellular H2O2 in cultured endothelial cells. Arch Biochem Biophys 302:348–355

    Article  CAS  PubMed  Google Scholar 

  21. Calera JA, Herman D, Calderone R (2000) Identification of Ypd1, a gene of Candida albicans which encodes a two-component phosphohistidine intermediate protein. Yeast 16:1053–1059

    Article  CAS  PubMed  Google Scholar 

  22. Gutteridge JM (1993) Anthracycline toxicity, iron and oxygen radicals, and chelation therapy. J Lab Clin Med 122:228–229

    CAS  PubMed  Google Scholar 

  23. Nordberg J, Arnér ES (2001) Reactive oxygen species, antioxidants, and the mammalian thioredoxin system. Free Radic Biol Med 31:1287–1312

    Article  CAS  PubMed  Google Scholar 

  24. Ketela T, Brown JL, Stewart RC, Bussey H (1998) Yeast Skn7p activity is modulated by the Sln1p-Ypd1p osmosensor and contributes to regulation of the HOG pathway. Mol Gen Genet 259:372–378

    Article  CAS  PubMed  Google Scholar 

  25. Hayashi M, Maeda T (2006) Activation of the HOG pathway upon cold stress in Saccharomyces cerevisiae. J Biochem 139:797–803

    Article  CAS  PubMed  Google Scholar 

  26. Panadero J, Pallotti C, Rodríguez-Vargas S, Randez-Gil F, Prieto JA (2006) A downshift in temperature activates the high osmolarity glycerol (HOG) pathway, which determines freeze tolerance in Saccharomyces cerevisiae. J Biol Chem 281:4638–4645

    Article  CAS  PubMed  Google Scholar 

  27. Freeman JA, Bassler BL (1999) Sequence and function of LuxU: a two-component phosphorelay protein that regulates quorum sensing in Vibrio harveyi. J Bacteriol 181:899–906

    CAS  PubMed  Google Scholar 

  28. Nguyen AN, Lee A, Place W, Shiozaki K (2000) Multistep phosphorelay proteins transmit oxidative stress signals to the fission yeast stress-activated protein kinase. Mol Biol Cell 11:1169–1181

    CAS  PubMed  Google Scholar 

  29. Barnes EA, Porter LA, Lenormand JL, Dellinger RW, Donoghue DJ (2003) Human Spy1 promotes survival of mammalian cells following DNA damage. Cancer Res 63:3701–3707

    CAS  PubMed  Google Scholar 

  30. Gastwirt RF, Slavin DA, McAndrew CW, Donoghue DJ (2006) Spy1 expression prevents normal cellular responses to DNA damage: inhibition of apoptosis and checkpoint activation. J Biol Chem 281:35425–35435

    Article  CAS  PubMed  Google Scholar 

  31. McAndrew CW, Gastwirt RF, Donoghue DJ (2009) The atypical CDK activator Spy1 regulates the intrinsic DNA damage response and is dependent upon p53 to inhibit apoptosis. Cell Cycle 8:66–75

    Article  CAS  PubMed  Google Scholar 

  32. Golipour A, Myers D, Seagroves T, Murphy D, Evan GI, Donoghue DJ, Moorehead RA, Porter LA (2008) The Spy1/RINGO family represents a novel mechanism regulating mammary growth and tumorigenesis. Cancer Res 68:3591–3600

    Article  CAS  PubMed  Google Scholar 

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Acknowledgements

This research was supported by Basic Science Research Program through the National Research Foundation of Korea (NRF) funded by the Ministry of Education, Science and Technology (2009-0072536). The authors wish to acknowledge Ms Sun-Young Ji for her technical assistance.

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Authors and Affiliations

  1. Division of Life Sciences and Research Institute of Life Sciences, College of Natural Sciences, Kangwon National University, 192-1 Hyoja-2-dong, Chuncheon, 200-701, Korea

    Min-Hee Kang & Chang-Jin Lim

  2. College of Pharmacy, Sookmyung Women’s University, 52 Hyochangwon-gil, Seoul, 140-742, Korea

    Hyun-Joo Jung & Eun-Hee Park

  3. Department of Life Science, Ewha Womans University, Seoul, 120-750, Korea

    Dong-Hoon Hyun

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  1. Min-Hee Kang
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  2. Hyun-Joo Jung
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  5. Chang-Jin Lim
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Correspondence to Chang-Jin Lim.

Additional information

The sequence reported in this paper has been deposited in the GenBank database with the accession number AL034352.

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Kang, MH., Jung, HJ., Hyun, DH. et al. Protective roles and Pap1-dependent regulation of the Schizosaccharomyces pombe spy1 gene under nitrosative and nutritional stresses. Mol Biol Rep 38, 1129–1136 (2011). https://doi.org/10.1007/s11033-010-0210-3

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