Institut de Biochimie et Génétique Cellulaires, Université de Bordeaux, 1 Rue Camille Saint-Saëns, 33077 Bordeaux, France.
Institute of Biochemistry and Biophysics, Polish Academy of Sciences, 02-106 Warsaw, Poland.
Int J Mol Sci. 2020 Jul 18;21(14):5083. doi: 10.3390/ijms21145083.
Probing the pathogenicity and functional consequences of mitochondrial DNA (mtDNA) mutations from patient's cells and tissues is difficult due to genetic heteroplasmy (co-existence of wild type and mutated mtDNA in cells), occurrence of numerous mtDNA polymorphisms, and absence of methods for genetically transforming human mitochondria. Owing to its good fermenting capacity that enables survival to loss-of-function mtDNA mutations, its amenability to mitochondrial genome manipulation, and lack of heteroplasmy, is an excellent model for studying and resolving the molecular bases of human diseases linked to mtDNA in a controlled genetic background. Using this model, we previously showed that a pathogenic mutation in mitochondrial gene (m.9191T>C), that converts a highly conserved leucine residue into proline in human ATP synthase subunit (L222P), severely compromises the assembly of yeast ATP synthase and reduces by 90% the rate of mitochondrial ATP synthesis. Herein, we report the isolation of intragenic suppressors of this mutation. In light of recently described high resolution structures of ATP synthase, the results indicate that the m.9191T>C mutation disrupts a four α-helix bundle in subunit and that the leucine residue it targets indirectly optimizes proton conduction through the membrane domain of ATP synthase.
由于遗传异质性（细胞中野生型和突变型 mtDNA 的共存）、大量 mtDNA 多态性的发生以及缺乏遗传转化人线粒体的方法，从患者的细胞和组织中探测线粒体 DNA（mtDNA）突变的致病性和功能后果是困难的。由于其良好的发酵能力，使其能够在功能丧失的 mtDNA 突变中存活，其易于进行线粒体基因组操作，以及缺乏异质性，是研究和解决与 mtDNA 相关的人类疾病的分子基础的理想模型，在受控遗传背景下。使用该模型，我们之前表明，线粒体基因（m.9191T>C）中的一个致病性突变，将人类 ATP 合酶亚基中的高度保守亮氨酸残基转换为脯氨酸（L222P），严重破坏酵母 ATP 合酶的组装，并使线粒体 ATP 合成的速率降低 90%。在此，我们报告了该突变的基因内抑制子的分离。鉴于最近描述的 ATP 合酶的高分辨率结构，结果表明 m.9191T>C 突变破坏了亚基中的一个四螺旋束，并且它靶向的亮氨酸残基间接优化了质子通过 ATP 合酶膜域的传导。
