Miyao A, Theeragool G, Takeuchi M, Kobayashi Y
Faculty of Agriculture, Tokyo University of Agriculture and Technology, Fuchu, Japan.
J Bacteriol. 1993 Jul;175(13):4081-6. doi: 10.1128/jb.175.13.4081-4086.1993.
Expression of the Bacillus subtilis sporulation gene spoVE was examined by runoff transcription assay with an RNA polymerase preparation obtained from vegetative and sporulating cells. Transcripts from tandem promoters (P1 and P2 promoters) located just upstream of the spoVE structure gene were detected. The transcription of spoVE initiated within an hour after the onset of sporulation and coincided with the presence of RNA polymerase associated with a 33-kDa protein. Amino acid sequence analysis of the 33-kDa protein revealed that it is a sigma factor, sigma E. Reconstitution analysis of sigma E purified from the sporulating cell extracts and vegetative core RNA polymerase showed that sigma E recognizes the P2 promoter. SpoVE protein could not be synthesized in the transcription-translation coupled system prepared from vegetative cells (M. Okamoto, S. Fukui, and Y. Kobayashi, Agric. Biol. Chem. 49:1077-1082, 1985). However, addition of sigma E-associated RNA polymerase to the coupled system restored SpoVE protein synthesis. These results indicate that spoVE expression in sporulating cells is controlled essentially by sigma E-associated RNA polymerase.
利用从营养细胞和芽孢形成细胞中获得的RNA聚合酶制剂，通过径流转录分析检测枯草芽孢杆菌芽孢形成基因spoVE的表达。检测到位于spoVE结构基因上游的串联启动子（P1和P2启动子）的转录本。spoVE的转录在芽孢形成开始后一小时内启动，并与与33 kDa蛋白相关的RNA聚合酶的存在同时发生。对33 kDa蛋白的氨基酸序列分析表明，它是一种sigma因子，即sigma E。从芽孢形成细胞提取物和营养核心RNA聚合酶中纯化的sigma E的重组分析表明，sigma E识别P2启动子。在从营养细胞制备的转录-翻译偶联系统中无法合成SpoVE蛋白（M.冈本、S.福井和Y.小林，《农业生物化学》49:1077 - 1082，1985）。然而，向偶联系统中添加与sigma E相关的RNA聚合酶可恢复SpoVE蛋白的合成。这些结果表明，芽孢形成细胞中spoVE的表达基本上由与sigma E相关的RNA聚合酶控制。
