Department of Biology, Tufts University, Medford, MA 02155.
Federal Reserve Bank of Atlanta, Atlanta, GA 30309.
Proc Natl Acad Sci U S A. 2021 Feb 2;118(5). doi: 10.1073/pnas.2013080118.
Nearly 50 hereditary diseases result from the inheritance of abnormally long repetitive DNA microsatellites. While it was originally believed that the size of inherited repeats is the key factor in disease development, it has become clear that somatic instability of these repeats throughout an individual's lifetime strongly contributes to disease onset and progression. Importantly, somatic instability is commonly observed in terminally differentiated, postmitotic cells, such as neurons. To unravel the mechanisms of repeat instability in nondividing cells, we created an experimental system to analyze the mutability of Friedreich's ataxia (GAA) repeats during chronological aging of quiescent Unexpectedly, we found that the predominant repeat-mediated mutation in nondividing cells is large-scale deletions encompassing parts, or the entirety, of the repeat and adjacent regions. These deletions are caused by breakage at the repeat mediated by mismatch repair (MMR) complexes MutSβ and MutLα and DNA endonuclease Rad1, followed by end-resection by Exo1 and repair of the resulting double-strand breaks (DSBs) via nonhomologous end joining. We also observed repeat-mediated gene conversions as a result of DSB repair via ectopic homologous recombination during chronological aging. Repeat expansions accrue during chronological aging as well-particularly in the absence of MMR-induced DSBs. These expansions depend on the processivity of DNA polymerase δ while being counteracted by Exo1 and MutSβ, implicating nick repair. Altogether, these findings show that the mechanisms and types of (GAA) repeat instability differ dramatically between dividing and nondividing cells, suggesting that distinct repeat-mediated mutations in terminally differentiated somatic cells might influence Friedreich's ataxia pathogenesis.
近 50 种遗传性疾病是由于异常长的重复 DNA 微卫星的遗传导致的。虽然最初认为遗传重复的大小是疾病发展的关键因素，但现在已经清楚，这些重复在个体一生中的体细胞不稳定性强烈促成疾病的发生和进展。重要的是，体细胞不稳定性通常在终末分化、有丝分裂后的细胞中观察到，如神经元。为了解析非分裂细胞中重复不稳定性的机制，我们创建了一个实验系统，以分析静止期细胞衰老过程中弗里德里希共济失调（GAA）重复的易变性。出乎意料的是，我们发现非分裂细胞中主要的重复介导突变是大规模缺失，包括重复及其相邻区域的部分或全部。这些缺失是由错配修复（MMR）复合物 MutSβ和 MutLα以及 DNA 内切酶 Rad1介导的重复断裂引起的，随后由 Exo1 进行末端切除，并通过非同源末端连接修复产生的双链断裂（DSB）。我们还观察到，在静止期细胞衰老过程中，通过异位同源重组进行 DSB 修复，导致重复介导的基因转换。重复扩展也会随着时间的推移而积累，特别是在没有 MMR 诱导的 DSB 的情况下。这些扩展依赖于 DNA 聚合酶 δ 的连续性，而 Exo1 和 MutSβ 则与之拮抗，暗示着缺口修复。总之，这些发现表明，在有丝分裂和非有丝分裂细胞之间，（GAA）重复不稳定性的机制和类型存在显著差异，这表明终末分化的体细胞中不同的重复介导突变可能影响弗里德里希共济失调的发病机制。
