Department of Biochemistry, Jichi Medical University, Shimotsuke, Tochigi 329-0498, Japan.
Exp Cell Res. 2011 Jan 15;317(2):210-20. doi: 10.1016/j.yexcr.2010.10.008. Epub 2010 Oct 15.
The segregation and transmission of the mitochondrial genome in humans are complicated processes but are particularly important for understanding the inheritance and clinical abnormalities of mitochondrial disorders. However, the molecular mechanism of the segregation of mitochondrial DNA (mtDNA) is largely unclear. In this study, we demonstrated that human mitochondrial transcription factor A (TFAM) is required for the segregation of mtDNA in cultured cells. RNAi-mediated knockdown of TFAM in HeLa cells resulted in the enlarged mtDNA, as indicated by the assembly of fluorescent signals stained with PicoGreen. Fluorescent in situ hybridization confirmed the enlarged mtDNA and further showed the existence of increased numbers of mitochondria lacking mtDNA signals in TFAM knockdown cells. By complementation analysis, the C-terminal tail of TFAM, which enhances its affinity with DNA, was found to be required for the appropriate distribution of mtDNA. Furthermore, we found that TFAM knockdown induced asymmetric segregation of mtDNA between dividing daughter cells. These results suggest an essential role for human TFAM in symmetric segregation of mtDNA.
线粒体基因组在人类中的分离和传递是复杂的过程，但对于理解线粒体疾病的遗传和临床异常尤为重要。然而，线粒体 DNA（mtDNA）分离的分子机制在很大程度上尚不清楚。在这项研究中，我们证明了人线粒体转录因子 A（TFAM）对于培养细胞中线粒体 DNA 的分离是必需的。用 RNAi 介导的 TFAM 在 HeLa 细胞中的敲低导致 mtDNA 增大，如用 PicoGreen 染色的荧光信号组装所表明的那样。荧光原位杂交进一步证实了 mtDNA 的增大，并显示在 TFAM 敲低细胞中存在更多缺乏 mtDNA 信号的线粒体。通过互补分析，发现 TFAM 的 C 末端尾巴增强了其与 DNA 的亲和力，对于 mtDNA 的适当分布是必需的。此外，我们发现 TFAM 敲低诱导 mtDNA 在分裂子细胞之间的不对称分离。这些结果表明人类 TFAM 在 mtDNA 的对称分离中起着重要作用。
