RNA 3′ polyadenylation is known to serve diverse purposes in biology, in particular, regulating mRNA stability and translation. Here we determined that, upon exposure to high levels of the intercalating agent ethidium bromide (EtBr), greater than those required to suppress mitochondrial transcription, mitochondrial tRNAs in human cells became polyadenylated. Relaxation of the inducing stress led to rapid turnover of the polyadenylated tRNAs. The extent, kinetics and duration of tRNA polyadenylation were EtBr dose-dependent, with mitochondrial tRNAs differentially sensitive to the stress. RNA interference and inhibitor studies indicated that ongoing mitochondrial ATP synthesis, plus the mitochondrial poly(A) polymerase and SUV3 helicase were required for tRNA polyadenylation, while polynucleotide phosphorylase counteracted the process and was needed, along with SUV3, for degradation of the polyadenylated tRNAs. Doxycycline treatment inhibited both tRNA polyadenylation and turnover, suggesting a possible involvement of the mitoribosome, although other translational inhibitors had only minor effects. The dysfunctional tRNA^Leu(UUR) bearing the pathological A3243G mutation was constitutively polyadenylated at a low level, but this was markedly enhanced after doxycycline treatment. We propose that polyadenylation of structurally and functionally abnormal mitochondrial tRNAs entrains their PNPase/SUV3-mediated destruction, and that this pathway could play an important role in mitochondrial diseases associated with tRNA mutations.

中文翻译：

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人细胞中结构异常的线粒体tRNA的聚腺苷酸化和降解

已知RNA 3'聚腺苷酸在生物学中具有多种用途，特别是调节mRNA的稳定性和翻译。在这里，我们确定，暴露于高水平的嵌入剂溴化乙锭（EtBr）大于抑制线粒体转录所需的嵌入量，人细胞中的线粒体tRNA变成了聚腺苷酸化。诱导应力的松弛导致聚腺苷酸化tRNA的快速周转。tRNA聚腺苷酸化的程度，动力学和持续时间是EtBr剂量依赖性的，线粒体tRNA对压力的敏感性不同。RNA干扰和抑制剂研究表明，正在进行的线粒体ATP合成，加上线粒体poly（A）聚合酶和SUV3解旋酶是tRNA聚腺苷酸化所必需的，而多核苷酸磷酸化酶可以抵消这一过程，因此需要与SUV3一起降解聚腺苷酸化的tRNA。强力霉素的治疗同时抑制了tRNA聚腺苷酸化和周转，这表明线粒体可能参与其中，尽管其他翻译抑制剂的作用很小。功能失调的tRNA带有病理性A3243G突变的^Leu（UUR）在低水平上组成型聚腺苷酸化，但在强力霉素处理后明显增强。我们建议结构和功能异常的线粒体tRNA的聚腺苷酸夹带其PNPase / SUV3介导的破坏，并且此途径可能在与tRNA突变相关的线粒体疾病中发挥重要作用。