Guan M X, Enriquez J A, Fischel-Ghodsian N, Puranam R S, Lin C P, Maw M A, Attardi G
Division of Biology, California Institute of Technology, Pasadena, California 91125, USA.
Mol Cell Biol. 1998 Oct;18(10):5868-79. doi: 10.1128/MCB.18.10.5868.
The pathogenetic mechanism of the deafness-associated mitochondrial DNA (mtDNA) T7445C mutation has been investigated in several lymphoblastoid cell lines from members of a New Zealand pedigree exhibiting the mutation in homoplasmic form and from control individuals. We show here that the mutation flanks the 3' end of the tRNASer(UCN) gene sequence and affects the rate but not the sites of processing of the tRNA precursor. This causes an average reduction of approximately 70% in the tRNASer(UCN) level and a decrease of approximately 45% in protein synthesis rate in the cell lines analyzed. The data show a sharp threshold in the capacity of tRNASer(UCN) to support the wild-type protein synthesis rate, which corresponds to approximately 40% of the control level of this tRNA. Strikingly, a 7445 mutation-associated marked reduction has been observed in the level of the mRNA for the NADH dehydrogenase (complex I) ND6 subunit gene, which is located approximately 7 kbp upstream and is cotranscribed with the tRNASer(UCN) gene, with strong evidence pointing to a mechanistic link with the tRNA precursor processing defect. Such reduction significantly affects the rate of synthesis of the ND6 subunit and plays a determinant role in the deafness-associated respiratory phenotype of the mutant cell lines. In particular, it accounts for their specific, very significant decrease in glutamate- or malate-dependent O2 consumption. Furthermore, several homoplasmic mtDNA mutations affecting subunits of NADH dehydrogenase may play a synergistic role in the establishment of the respiratory phenotype of the mutant cells.
在来自一个新西兰家系成员的多个淋巴母细胞系中，已对与耳聋相关的线粒体DNA（mtDNA）T7445C突变的致病机制进行了研究，这些家系成员呈现纯质形式的突变，同时也对对照个体进行了研究。我们在此表明，该突变位于tRNASer(UCN)基因序列的3'端侧翼，影响tRNA前体的加工速率，但不影响加工位点。这导致在分析的细胞系中，tRNASer(UCN)水平平均降低约70%，蛋白质合成速率降低约45%。数据显示，tRNASer(UCN)支持野生型蛋白质合成速率的能力存在一个明显的阈值，该阈值约为该tRNA对照水平的40%。令人惊讶的是，在位于上游约7 kbp处且与tRNASer(UCN)基因共转录的NADH脱氢酶（复合体I）ND6亚基基因的mRNA水平上，观察到与7445突变相关的显著降低，有力证据表明其与tRNA前体加工缺陷存在机制上的联系。这种降低显著影响ND6亚基的合成速率，并在突变细胞系与耳聋相关的呼吸表型中起决定性作用。特别是，它解释了它们在依赖谷氨酸或苹果酸的氧气消耗方面的特定且非常显著的降低。此外，几种影响NADH脱氢酶亚基的纯质mtDNA突变可能在突变细胞呼吸表型的形成中起协同作用。
