The radiation responsive promoter of the Deinococcus radiodurans pprA gene
The radiation responsive promoter of the Deinococcus radiodurans pprA gene
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DOI：
10.1016/j.gene.2005.07.035
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发表时间：
2005-12-19
期刊：
影响因子：
3.5
通讯作者：
Narumi, I
中科院分区：
文献类型：
--
作者：
Ohba, H;Satoh, K;Narumi, I
在先前的研究中，我们鉴定了一种新的辐射诱导蛋白PprA，其在耐辐射异常球菌的辐射抗性中起关键作用[Narumi，I.，Satoh，K.，美国，Cui，S.，Funayama，T.，Kitayama，S.，渡边，H.，2004. PprA：一种来自耐辐射球菌的新蛋白，可刺激DNA连接。摩尔Microbiol. 54，278-285.]。尽管对D.在抗辐射蛋白中，对辐射诱导蛋白的辐射响应启动子知之甚少。在这项研究中，三个转录起始点的pprA mRNA的引物延伸分析确定，位于位置-156，-154和-22上游的pprA翻译起始位点。在暴露于2kGy的细胞中，三种延伸产物的量增加，然后孵育后0.5小时。这表明存在至少两个辐射响应性启动子用于pprA表达。使用荧光素酶报告基因测定对pprA基因上游区域的功能表征揭示，远端启动子位于距离翻译起始位点的位置-208和-156之间，而近端启动子位于位置-57和-22之间。位于位置-57和-38之间的区域对于近端启动子活性是必不可少的。位于-33位的胸腺嘧啶的定点突变导致启动子活性的严重损害，并表明胸腺嘧啶作为近端辐射响应启动子的主碱基起作用。D.耐辐射菌ppr 1基因被认为是辐射反应中的一般开关[Hua，Y.，成海岛高，G.，Tian，B.，Satoh，K.，Kitayama，S.，Shen，B.，2003. PprI：一个负责耐辐射球菌极端辐射抗性的通用开关。生物化学生物物理学通信资源306，354-360.]。我们研究了ppr 1中断对pprA启动子活性的影响。结果表明，上调pprA表达的pprI基因产物是在启动子水平触发。(c)2005 Elsevier B. V.保留所有权利。
In a previous study, we identified a novel radiation-inducible protein PprA that plays a critical role in the radiation resistance of Deinococcus radiodurans [Narumi, I., Satoh, K., Cui, S., Funayama, T., Kitayama, S., Watanabe, H., 2004. PprA: a novel protein from Deinococcus radiodurans that stimulates DNA ligation. Mol. Microbiol. 54, 278-285.]. Despite the interest in mechanisms underlying radiation responses in D. radiodurans, little is known about the radiation responsive promoter for radiation-inducible proteins. In this study, three transcriptional start points for pprA mRNA were identified by primer extension analysis, located at positions -156, -154 and -22 upstream from the pprA translation initiation site. The amount of the three extended products increased in cells exposed to 2 kGy followed by a 0.5-h post-incubation. This suggested the existence of at least two radiation responsive promoters for pprA expression. Functional characterization of the upstream region of the pprA gene using a luciferase reporter assay revealed that the distal promoter is located between positions -208 and -156 from the translation initiation site, while the proximal promoter is located between positions -57 and -22. The region located between positions -57 and -38 was indispensable for proximal promoter activity. Site-directed mutagenesis of a thymine positioned at -33 resulted in severe impairment of promoter activity, and suggested that the thymine functions as a master base for the proximal radiation responsive promoter. The product of the D. radiodurans ppr1 gene is thought to be a general switch in the radiation response [Hua, Y., Narumi, I., Gao, G., Tian, B., Satoh, K., Kitayama, S., Shen, B., 2003. PprI: a general switch responsible for extreme radioresistance of Deinococcus radiodurans. Biochem. Biophys. Res. Commun. 306, 354-360.]. We examined the effect of ppr1 disruption on pprA promoter activity. The results suggested that up-regulation of pprA expression by the pprI gene product is triggered at the promoter level. (c) 2005 Elsevier B.V. All rights reserved.