Characterization and regulation of the gene encoding monothiol glutaredoxin 3 in the fission yeast Schizosaccharomyces pombe.
Glutaredoxins (Grxs) are thioloxidoreductases which are required for maintaining thiol/disulfide equilibrium in living cells. The Grx3 gene, which encodes one of the three monothiol Grxs in the fission yeast Schizosaccharomyces pombe, was characterized, and its transcriptional regulation studied. Genomic DNA encoding Grx3 was isolated by PCR, and a plasmid pTT3 carrying this DNA was produced. The DNA sequence has 1,267 bp, which would encode a monothiol Grx of 166 amino acids with a molecular mass of 18.3 kDa. The putative protein has 27% homology with Grx5, and contains many hydrophobic amino acid residues in its N-terminal region. S. pombe cells harboring pTT3 had increased Grx activity and enhanced survival on minimal medium plates containing aluminum (5 mM), BSO (0.05 mM), menadione (0.01 mM) or cadmium (0.2 mM). The 568 bp upstream region of Grx3 was fused into the promoterless beta-galactosidase gene of the shuttle vector YEp367R to generate fusion plasmid pMJS10. Potassium chloride (KCl) and metals including aluminum and cadmium enhanced the synthesis of beta-galactosidase from the fusion gene. The synthesis of beta-galactosidase was also enhanced, in a Pap1-dependent manner, by fermentable carbon sources such as glucose (at low concentrations) and sucrose, but not by non-fermentable carbon sources such as ethanol and acetate. Grx3 mRNA increased in response to treatment with BSO. These observations indicate that S. pombe Grx3 is involved in the response to stress, and is regulated by stress.
Moon
Jeong-Su
JS
Division of Life Sciences, College of Natural Sciences, Kangwon National University, Chuncheon 200-701, Korea.
Lim
Hye-Won
HW
Park
Eun-Hee
EH
Lim
Chang-Jin
CJ
D000595:Amino Acid Sequence; D001483:Base Sequence; D050976:Basic-Leucine Zipper Transcription Factors / Q000378:metabolism; D019328:Buthionine Sulfoximine / Q000378:metabolism / Q000494:pharmacology; D015966:Gene Expression Regulation, Fungal; D054477:Glutaredoxins; D008670:Metals / Q000378:metabolism / Q000494:pharmacology; D008969:Molecular Sequence Data; D009997:Osmotic Pressure; D018384:Oxidative Stress / Q000235:genetics / Q000502:physiology; D010088:Oxidoreductases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D000073718:Pancreatitis-Associated Proteins; D012568:Schizosaccharomyces / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D029702:Schizosaccharomyces pombe Proteins / Q000235:genetics / Q000378:metabolism; D013997:Time Factors; D001616:beta-Galactosidase / Q000235:genetics / Q000378:metabolism
D050976:Basic-Leucine Zipper Transcription Factors; D054477:Glutaredoxins; D008670:Metals; D000073718:Pancreatitis-Associated Proteins; C105096:Pap1 protein, S pombe; C000612141:REG3A protein, human; D029702:Schizosaccharomyces pombe Proteins; D019328:Buthionine Sulfoximine; D010088:Oxidoreductases; D001616:beta-Galactosidase