Takata K, Yoshida H, Hirose F, Yamaguchi M, Kai M, Oshige M, Sakimoto I, Koiwai O, Sakaguchi K
Department of Applied Biological Science, Faculty of Science and Technology, Science University of Tokyo, 2641 Yamazaki, Noda-shi, Chiba-ken 278-8510, Japan.
Biochem Biophys Res Commun. 2001 Sep 21;287(2):474-83. doi: 10.1006/bbrc.2001.5528.
We cloned a cDNA for Drosophila mitochondrial transcription factor A (D-mtTFA) and characterized the recombinant protein. In Drosophila Kc cells, D-mtTFA was localized in the mitochondria, but not in the nucleus. By repetitive precipitation with His-tag and PCR amplification, the consensus nucleotide sequence for D-mtTFA-binding was determined to be 5'-TTATC/G. The binding sequence was found to be clustered in the A + T region of mitochondrial DNA which is suggested to be a replication origin and promoter region for light strand and heavy strand. We found a DNA replication-related element (DRE)-like sequence located upstream of the transcription initiation site of the D-mtTFA gene and obtained results indicating that DRE-binding factor (DREF) can bind to the DRE-like sequence of the D-mtTFA gene. The data suggest that transcription of the D-mtTFA gene is under control of the DRE/DREF regulatory system. Based on these results, the functions of D-mtTFA were discussed in relation to mitochondrial biogenesis of Drosophila melanogaster.
我们克隆了果蝇线粒体转录因子A（D-mtTFA）的cDNA并对重组蛋白进行了表征。在果蝇Kc细胞中，D-mtTFA定位于线粒体而非细胞核。通过His标签的重复沉淀和PCR扩增，确定D-mtTFA结合的共有核苷酸序列为5'-TTATC/G。发现该结合序列聚集在线粒体DNA的A + T区域，该区域被认为是轻链和重链的复制起点和启动子区域。我们在D-mtTFA基因转录起始位点上游发现了一个类似DNA复制相关元件（DRE）的序列，并获得了表明DRE结合因子（DREF）可与D-mtTFA基因的DRE样序列结合的结果。数据表明D-mtTFA基因的转录受DRE/DREF调节系统的控制。基于这些结果，讨论了D-mtTFA的功能与黑腹果蝇线粒体生物发生的关系。
