Friedreich's ataxia (FRDA) is a human hereditary disease caused by the presence of expanded (GAA)n repeats in the first intron of the FXN gene [V. Campuzano et al., Science 271, 1423-1427 (1996)]. In somatic tissues of FRDA patients, (GAA)n repeat tracts are highly unstable, with contractions more common than expansions [R. Sharma et al., Hum. Mol. Genet. 11, 2175-2187 (2002)]. Here we describe an experimental system to characterize GAA repeat contractions in yeast and to conduct a genetic analysis of this process. We found that large-scale contraction is a one-step process, resulting in a median loss of ∼60 triplet repeats. Our genetic analysis revealed that contractions occur during DNA replication, rather than by various DNA repair pathways. Repeats contract in the course of lagging-strand synthesis: The processivity subunit of DNA polymerase δ, Pol32, and the catalytic domain of Rev1, a translesion polymerase, act together in the same pathway to counteract contractions. Accumulation of single-stranded DNA (ssDNA) in the lagging-strand template greatly increases the probability that (GAA)n repeats contract, which in turn promotes repeat instability in rfa1, rad27, and dna2 mutants. Finally, by comparing contraction rates for homopurine-homopyrimidine repeats differing in their mirror symmetry, we found that contractions depend on a repeat's triplex-forming ability. We propose that accumulation of ssDNA in the lagging-strand template fosters the formation of a triplex between the nascent and fold-back template strands of the repeat. Occasional jumps of DNA polymerase through this triplex hurdle, result in repeat contractions in the nascent lagging strand.

中文翻译：

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DNA复制过程中，由于弗里德赖希共济失调GAA重复序列在酵母中发生大规模收缩，这是因为它们具有三链体形成能力。

弗里德赖希共济失调（FRDA）是一种人类遗传病，是由FXN基因的第一个内含子中存在扩展（GAA）n重复序列引起的。Campuzano等，Science 271，1423-1427（1996）]。在FRDA患者的身体组织中，（GAA）n重复道非常不稳定，收缩比扩张更常见。Sharma et al。，Hum。大声笑 基因 11，2175-2187（2002）]。在这里，我们描述了一个表征酵母中GAA重复收缩并对该过程进行遗传分析的实验系统。我们发现大规模收缩是一个一步的过程，导致中位数损失约60个三联体重复。我们的遗传分析表明，收缩是在DNA复制过程中发生的，而不是通过各种DNA修复途径发生的。在滞后链合成过程中重复收缩：DNA聚合酶δ，Pol32和Rev1的催化域（一种跨病变的聚合酶）的合成亚基以相同的途径共同作用以抵消收缩。滞后链模板中单链DNA（ssDNA）的积累大大增加了（GAA）n重复收缩的可能性，这反过来又促进了rfa1，rad27和dna2突变体的重复不稳定。最后，通过比较不同镜象对称性的高嘌呤-高嘧啶重复序列的收缩率，我们发现收缩取决于重复序列的三链体形成能力。我们提出，ssDNA在滞后链模板中的积累促进了重复序列的新生和折返模板链之间三链体的形成。DNA聚合酶偶尔会通过这个三重障碍而跳跃，