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Molecular cloning, characterization and regulation of a peroxiredoxin gene from Schizosaccharomyces pombe

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Abstract

A gene encoding a putative peroxiredoxin (Prx) of the fission yeast Schizosaccharomyces pombe was characterized and its regulation was studied. The full length of the prx gene was introduced into the shuttle vector pRS316 after PCR amplification, resulting in the recombinant plasmid pPrx10. The determined DNA sequence carries 1,327 bp encoding a putative Prx with a molecular mass of 19,510 Da. Prx activity was significantly increased in the S. pombe cells harboring pPrx10. The accelerated growth was observed in the S. pombe/pPrx10 cells, implying the involvement of the cloned gene in the yeast growth. To study transcriptional regulation of the prx gene, the prx-lacZ fusion gene was constructed using the yeast-E. coli shuttle vector YEp367R, and named pPrxup10. The synthesis of β-galactosidase from the fusion gene was enhanced under carbon source-limited conditions and nitrogen starvation. Under the same growth conditions, the prx mRNA levels of the wild-type yeast cells were increased. The prx mRNA level was markedly decreased in the Pap1-negative mutant, compared with that in the wild-type yeast, suggesting that the basal expression of the prx gene is mediated by a transcription factor, Pap1. The reactive oxygen species (ROS) level was diminished in the S. pombe/pPrx10 cells than in the control cells. The extra copies of the prx gene were able to resist elevation of ROS level under limited carbon source condition and menadione treatment. In brief, the S. pombe Prx is linked with the yeast growth and up-regulated by metabolic oxidative stress on a transcriptional level. The Prx protein is partly responsible for maintaining low ROS level under normal and stressful growth conditions in the fission yeast.

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Abbreviations

ONPG:

ο-Nitrophenyl-β-d-galactoside

Prx:

Peroxiredoxin

PCR:

Polymerase chain reaction

ROS:

Reactive oxygen species

RT-PCR:

Reverse transcriptase-polymerase chain reaction

S. pombe :

Schizosaccharomyces pombe

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Acknowledgements

This work was supported by the SRC program of MOST/KOSEF (R11-2005-017).

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Authors and Affiliations

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

    Ga-Young Kang & Chang-Jin Lim

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

    Eun-Hee Park

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  1. Ga-Young Kang
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Correspondence to Eun-Hee Park or Chang-Jin Lim.

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Kang, GY., Park, EH. & Lim, CJ. Molecular cloning, characterization and regulation of a peroxiredoxin gene from Schizosaccharomyces pombe . Mol Biol Rep 35, 387–395 (2008). https://doi.org/10.1007/s11033-007-9098-y

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