Rearrangements of Human Mitochondrial DNA (mtDNA): New Insights into the Regulation of mtDNA Copy Number and Gene Expression

Mitochondria from patients with Kearns–Sayre syndrome harboring large-scale rearrangements of human mitochondrial DNA (mtDNA; both partial deletions and a partial duplication) were introduced into human cells lacking endogenous mtDNA. Cytoplasmic hybrids containing 100% wild-type mtDNA, 100% mtDNA with partial duplications, and 100% mtDNA with partial deletions were isolated and characterized. The cell lines with 100% deleted mtDNAs exhibited a complete impairment of respiratory chain function and oxidative phosphorylation. In contrast, there were no detectable respiratory chain or protein synthesis defects in the cell lines with 100% duplicated mtDNAs. Unexpectedly, the mass of mtDNA was identical in all cell lines, despite the fact that different lines contained mtDNAs of vastly different sizes and with different numbers of replication origins, suggesting that mtDNA copy number may be regulated by tightly controlled mitochondrial dNTP pools. In addition, quantitation of mtDNA-encoded RNAs and polypeptides in these lines provided evidence that mtDNA gene copy number affects gene expression, which, in turn, is regulated at both the post-transcriptional and translational levels.

将携带人类线粒体DNA（mtDNA）大规模重排（涵盖部分缺失与部分重复两类变异）的卡恩斯-塞尔综合征（Kearns–Sayre syndrome）患者来源的线粒体，导入至不含内源性线粒体DNA的人类细胞中。随后分离并鉴定了三类胞质杂种：分别包含100%野生型mtDNA、100%携带部分重复的mtDNA，以及100%携带部分缺失的mtDNA。携带100%缺失型mtDNA的细胞系完全丧失了呼吸链功能与氧化磷酸化能力。与之相对，携带100%重复型mtDNA的细胞系未检测到呼吸链或蛋白质合成相关缺陷。出乎意料的是，尽管不同细胞系所含mtDNA的分子大小差异显著且复制起始位点数量各不相同，但所有细胞系的mtDNA总含量完全一致，这提示线粒体DNA拷贝数或受严格调控的线粒体脱氧核苷三磷酸（dNTP）池调控。此外，对这些细胞系中mtDNA编码的RNA与多肽进行定量分析后证实，mtDNA基因拷贝数可影响基因表达，而基因表达的调控同时发生在转录后与翻译两个水平。