Kang Ga-Young, Park Eun-Hee, Lim Chang-Jin
Division of Life Sciences, College of Natural Sciences, Kangwon National University, 192-1 Hyoja-2-dong, Chuncheon, Korea.
Mol Biol Rep. 2008 Sep;35(3):387-95. doi: 10.1007/s11033-007-9098-y. Epub 2007 May 26.
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 beta-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.
对裂殖酵母粟酒裂殖酵母中一个假定的过氧化物还原酶（Prx）编码基因进行了表征，并研究了其调控机制。prx基因全长经PCR扩增后被导入穿梭载体pRS316，得到重组质粒pPrx10。测定的DNA序列包含1327 bp，编码一个假定的Prx，分子量为19510 Da。在携带pPrx10的粟酒裂殖酵母细胞中，Prx活性显著增加。在粟酒裂殖酵母/pPrx10细胞中观察到生长加速，这意味着克隆基因参与了酵母生长。为了研究prx基因的转录调控，使用酵母-大肠杆菌穿梭载体YEp367R构建了prx-lacZ融合基因，并命名为pPrxup10。在碳源受限条件和氮饥饿情况下，融合基因的β-半乳糖苷酶合成增强。在相同生长条件下，野生型酵母细胞的prx mRNA水平升高。与野生型酵母相比，Pap1阴性突变体中的prx mRNA水平显著降低，这表明prx基因的基础表达由转录因子Pap1介导。粟酒裂殖酵母/pPrx10细胞中的活性氧（ROS）水平低于对照细胞。prx基因的额外拷贝能够在有限碳源条件和甲萘醌处理下抵抗ROS水平的升高。简而言之，粟酒裂殖酵母Prx与酵母生长相关，并在转录水平上受到代谢氧化应激的上调。Prx蛋白在裂殖酵母正常和应激生长条件下部分负责维持低ROS水平。
