He Xiangyu, Zhu Xiaoyu, Wang Xuexiang, Wang Wei, Dai Yu, Yan Qingfeng
College of Life Science, Zhejiang University, Hangzhou, Zhejiang, China.
PLoS One. 2013 Dec 10;8(12):e81490. doi: 10.1371/journal.pone.0081490. eCollection 2013.
The phenotypic manifestations of mitochondrial DNA (mtDNA) mutations are modulated by mitochondrial DNA haplotypes, nuclear modifier genes and environmental factors. The yeast mitochondrial 15S rRNA C1477G (P(R) or P(R) 454) mutation corresponds to the human 12S rRNA C1494T and A1555G mutations, which are well known as primary factors for aminoglycoside-induced nonsyndromic deafness. Here we report that the deletion of the nuclear modifier gene MTO2 suppressed the aminoglycoside-sensitivity of mitochondrial 15S rRNA C1477G mutation in Saccharomyces cerevisiae. First, the strain with a single mtDNA C1477G mutation exhibited hypersensitivity to neomycin. Functional assays indicated that the steady-state transcription level of mitochondrial DNA, the mitochondrial respiratory rate, and the membrane potential decreased significantly after neomycin treatment. The impaired mitochondria could not produce sufficient energy to maintain cell viability. Second, when the mto2 null and the mitochondrial C1477G mutations co-existed (mto2(P(R))), the oxygen consumption rate in the double mutant decreased markedly compared to that of the control strains (MTO2(P(S)), mto2(P(S)) and MTO2(P(R))). The expression levels of the key glycolytic genes HXK2, PFK1 and PYK1 in the mto2(P(R)) strain were stimulated by neomycin and up-regulated by 89%, 112% and 55%, respectively. The enhanced glycolysis compensated for the respiratory energy deficits, and could be inhibited by the glycolytic enzyme inhibitor. Our findings in yeast will provide a new insight into the pathogenesis of human deafness.
线粒体DNA（mtDNA）突变的表型表现受到线粒体DNA单倍型、核修饰基因和环境因素的调节。酵母线粒体15S rRNA C1477G（P(R)或P(R) 454）突变对应于人类12S rRNA C1494T和A1555G突变，这两个突变是氨基糖苷类药物诱导的非综合征性耳聋的主要因素。在此我们报道，核修饰基因MTO2的缺失抑制了酿酒酵母中线粒体15S rRNA C1477G突变对氨基糖苷类药物的敏感性。首先，具有单个mtDNA C1477G突变的菌株对新霉素表现出超敏感性。功能分析表明，新霉素处理后，线粒体DNA的稳态转录水平、线粒体呼吸速率和膜电位显著下降。受损的线粒体无法产生足够的能量来维持细胞活力。其次，当mto2缺失与线粒体C1477G突变共存（mto2(P(R))）时，与对照菌株（MTO2(P(S))、mto2(P(S))和MTO2(P(R))）相比，双突变体中的氧消耗率显著降低。新霉素刺激了mto2(P(R))菌株中关键糖酵解基因HXK2、PFK1和PYK1的表达水平，分别上调了89%、112%和55%。增强的糖酵解弥补了呼吸能量的不足，并且可以被糖酵解酶抑制剂抑制。我们在酵母中的发现将为人类耳聋的发病机制提供新的见解。
