Mechanisms of mistranslation-mediated mutator response

描述（由申请人提供）：突变在癌症发生和遗传性疾病以及微生物发病机制和耐药性中起关键作用。由于突变也会产生进化所需的遗传变异性，因此细胞不仅具有在正常生长条件下避免过度突变的机制，而且还具有响应环境和生理应激而促进突变的机制。在这个提议中，我们将研究最近发现的诱变途径，令人惊讶的是，该途径被发现是由编码tRNA的基因中的突变（mutA）触发的。 我们在上一个资助期的调查导致了几个重要的发现：蛋白质合成中的遗传缺陷引起的误译，或暴露于抗生素链霉素，可以诱导这种诱变途径，命名为TSM翻译应激诱导的诱变。TSM不同于众所周知的SOS诱变途径，并且似乎在某些条件下需要同源重组功能。TSM似乎是由修改后的形式的DMA聚合酶III的易错DMA合成介导的。这些发现意味着翻译和DMA复制之间存在以前未描述的联系，并提高了对细胞存活，适应，癌症发生和衰老具有重要意义的增变基因表型的可能性，这些表型可以通过比以前认识到的更大的靶基因库中的缺陷而产生。通过抗生素暴露和潜在地通过氨基酸饥饿诱导该途径表明环境和生理应激也可以通过该途径增加诱变。我们的工作进一步表明，氨基糖苷类抗生素通过这种以前未被认识的机制造成致突变风险。 在这里，我们建议通过以下三个特定的目的，研究误译导致这种增变表型的机制。(1)检验复制型DNA聚合酶的直接误译导致TSM表型的假设。(2)检验在Rec-mutA细胞中存在对经历偶发性异常复制周期的细胞的选择性杀伤的假设。(3)检验TSM表型是由偶发性超诱变循环引起的假设。

DESCRIPTION (provided by applicant): Mutations play critical roles in cancer initiation and genetic disorders, as well as in microbial pathogenesis and resistance to drugs. Because mutations also generate genetic variability that is required for evolution, cells have mechanisms not only for avoiding excess mutation during normal growth conditions, but also for promoting mutations in response to environmental and physiological stress. In this proposal we will investigate of a recently discovered pathway for mutagenesis that was found to be triggered, surprisingly, by a mutation (mutA) in a gene coding for a tRNA. Our investigation in the previous funding period has led to several important discoveries: mistranslation induced by genetic defects in protein synthesis, or exposure to the antibiotic streptomycin, can induce this mutagenic pathway, named TSM for translational stress-induced mutagenesis. TSM is distinct from the well known SOS mutagenesis pathway, and appears to require homologous recombination functions under certain conditions. TSM appears to be mediated by error-prone DMA synthesis by a modified form of DMA polymerase III. These findings imply the existence of previously undescribed links between translation and DMA replication, and raise the possibility that mutator phenotypes with significance for cell survival, adaptation, cancer initiation, and aging can arise through defects in a much larger repertoire of target genes than previously recognized. The induction of this pathway by antibiotic exposure, and potentially by amino acid starvation suggest that environmental and physiological stress can also increase mutagenesis by this pathway. Our work further shows that aminoglycoside antibiotics pose a mutagenic risk through this previously unrecognized mechanism. Here, we propose to investigate the mechanisms by which mistranslation leads this mutator phenotype through the following three Specific Aims. (1) Test the hypothesis that direct mistranslation of a replicative DMA polymerase leads to the TSM phenotype. (2) Test the hypothesis that in Rec- mutA cells there is selective killing of cells undergoing episodic aberrant replication cycles. (3) Test the hypothesis that the TSM phenotype results from episodic hyper-mutagenesis cycles.