Aberrant Double-Strand Break Repair Resulting in Half Crossovers in Mutants Defective for Rad51 or the DNA Polymerase δ Complex
Aberrant Double-Strand Break Repair Resulting in Half Crossovers in Mutants Defective for Rad51 or the DNA Polymerase δ Complex
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DOI：
10.1128/mcb.01469-08
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发表时间：
2009-03-15
影响因子：
5.3
通讯作者：
Symington, Lorraine S.
中科院分区：
文献类型：
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作者：
Smith, Catherine E.;Lam, Alicia F.;Symington, Lorraine S.
同源重组是DNA双链断裂（DSB）修复的一种无错机制。大多数DSB修复事件通过基因转换发生，限制修复位点下游标记的杂合性丢失（洛）并限制有害的染色体重排。只有一个末端可用于修复的DSB经历链侵入同源双链DNA，然后复制到染色体末端（断裂诱导复制[BIR]），导致链侵入位点下游的所有标记物的洛。使用基于转化的测定系统，我们表明，大多数明显的BIR事件中出现的二倍体酿酒酵母rad 51三角洲突变体是由于半交叉，而不是BIR。这些事件导致广泛的洛缺失，因为染色体III的一个臂被删除。在pol 32 Delta和pol 3-ct突变体中也观察到这种结果，这些突变体对DNA聚合酶delta（Pol delta）复合物的组分有缺陷。在Pol δ复合物突变体中形成的半交叉显示出有限的同源依赖性DNA合成的证据，并且部分依赖于Mus 81，这表明发生链侵入并且停滞的中间体随后被切割。与rad 51 δ突变体相反，Pol δ复合物突变体精通通过基因转换修复238-bp缺口。因此，对于rad 51突变体观察到的BIR缺陷是由于链侵入失败，而Pol δ复合物突变体精通链侵入，但不能完成重组起始的DNA合成的广泛区域。
Homologous recombination is an error-free mechanism for the repair of DNA double-strand breaks (DSBs). Most DSB repair events occur by gene conversion limiting loss of heterozygosity (LOH) for markers downstream of the site of repair and restricting deleterious chromosome rearrangements. DSBs with only one end available for repair undergo strand invasion into a homologous duplex DNA, followed by replication to the chromosome end (break-induced replication [BIR]), leading to LOH for all markers downstream of the site of strand invasion. Using a transformation-based assay system, we show that most of the apparent BIR events that arise in diploid Saccharomyces cerevisiae rad51 Delta mutants are due to half crossovers instead of BIR. These events lead to extensive LOH because one arm of chromosome III is deleted. This outcome is also observed in pol32 Delta and pol3-ct mutants, defective for components of the DNA polymerase delta (Pol delta) complex. The half crossovers formed in Pol delta complex mutants show evidence of limited homology-dependent DNA synthesis and are partially Mus81 dependent, suggesting that strand invasion occurs and the stalled intermediate is subsequently cleaved. In contrast to rad51 Delta mutants, the Pol delta complex mutants are proficient for repair of a 238-bp gap by gene conversion. Thus, the BIR defect observed for rad51 mutants is due to strand invasion failure, whereas the Pol delta complex mutants are proficient for strand invasion but unable to complete extensive tracts of recombination-initiated DNA synthesis.