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Stress-dependent regulation of a monothiol glutaredoxin gene from Schizosaccharomyces pombe

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Abstract:

Glutaredoxin (Grx) is a small, heat-stable protein acting as a multi-functional glutathione-dependent disulfide oxidoreductase. In this work, a gene encoding the monothiol glutaredoxin Grx4 was cloned from the genomic DNA of the fission yeast Schizosaccharomyces pombe. The determined DNA sequence carries 1706 bp, which is able to encode the putative 244 amino acid sequence of Grx with 27 099 Da. It does not contain an intron, and the sequence CGFS is found in the active site. Grx activity was increased 1.46-fold in S. pombe cells harboring the cloned Grx4 gene, indicating that the Grx4 gene is in vivo functioning. Although aluminum, cadmium, and hydrogen peroxide marginally enhanced the synthesis of -galactosidase from the Grx4-lacZ fusion gene, NO-generating sodium nitroprusside (0.5 mmol/L and 1.0 mmol/L) and potassium chloride (0.2 mol/L and 0.5 mol/L) significantly enhanced it. The Grx4 mRNA level was also enhanced after the treatment with sodium nitroprusside and potassium chloride. The synthesis of -galactosidase from the Grx4-lacZ gene was increased by fermentable carbon sources, such as glucose (lower than 2%) and sucrose, but not by nonfermentable carbon sources such as acetate and ethanol. The basal expression of the S. pombe Grx4 gene did not depend on the presence of Pap1. These results imply that the S. pombe monothiol Grx4 gene is genuinely functional and regulated by a variety of stresses.Key words: monothiol glutaredoxin, Pap1, regulation, Schizosaccharomyces pombe, stress response.

La glutarédoxine (Grx) est une petite protéine thermostable qui agit en tant que disulfure oxydoréductase multifonctionnelle dépendante du glutathion. Dans ces travaux, un gène codant une glutarédoxine de type monothiol Grx4 fut cloné à partir de l'ADN génomique de la levure scissipare Schizosaccharomyces pombe. La séquence déterminée d'ADN est d'une longueur de 1706 pb et et capable de coder la séquence putative de 244 acides aminés de Grx d'une masse moléculaire de 27 099 Da. Il ne contient pas d'intron et la séquence CGFS serait trop au site actif. L'activité de Grx fut augmentée d'un facteur de 1,46 chez des cellules de S. pombe contenant le gène Grx4 cloné, ce qui indique que le gène Grx4 est fonctionnel in vivo. Bien que l'aluminium, le cadmium et le peroxyde d'hydrogène ont marginalement stimulé la synthèse de la -galactosidase du gène de fusion Grx4-lacZ, le nitroprussure de sodium (0,5 mmol/L et 1,0 mmol/L) et le chlorure potassium (0,2 mol/L et 0,5 mol/L) l'ont augmenté significativement. Le niveau d'ARNm de Grx4 s'est également accru suite au traitement avec le nitroprussure de sodium et le chlorure de potassium. La synthèse de la -galactosidase à partir du gène Grx4-lacZ fut intensifiée par la présence de sources de carbone fermentable tel que le glucose (moins de 2 %) et le sucrose mais non par des sources de carbone non-fermentables tel que l'acétate et l'éthanol. L'expression basale du gène Grx4 de S. pombe n'était pas dépendante de la présence de Pap1. Ces résultats laissent entendre que le gène Grx4 de type monothiol de S. pombe est véritablement fonctionnel et sous le contrôle d'une variété de stress.Mots clés : monothiol glutarédoxine, Pap1, régulation, Schizosaccharomyces pombe, réponse au stress.[Traduit par la Rédaction]

Document Type: Research Article

Publication date: July 1, 2005

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  • Published since 1954, this monthly journal contains new research in the field of microbiology including applied microbiology and biotechnology; microbial structure and function; fungi and other eucaryotic protists; infection and immunity; microbial ecology; physiology, metabolism and enzymology; and virology, genetics, and molecular biology. It also publishes review articles and notes on an occasional basis, contributed by recognized scientists worldwide.
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