Abstract：
Although gene conversion (GC) in Saccharomyces cerevisiae is the most error-free way to repair double-strand breaks (DSBs), the mutation rate during homologous recombination is 1,000 times greater than during replication. Many mutations involve dissociating a partially copied strand from its repair template and re-aligning with the same or another template, leading to -1 frameshifts in homonucleotide runs, quasipalindrome (QP)-associated mutations and microhomology-mediated interchromosomal template switches. We studied GC induced by HO endonuclease cleavage at MATα, repaired by an HMR::KI-URA3 donor. We inserted into HMR::KI-URA3 an 18-bp inverted repeat where one arm had a 4-bp insertion. Most GCs yield MAT::KI-ura3::QP + 4 (Ura-) outcomes, but template-switching produces Ura+ colonies, losing the 4-bp insertion. If the QP arm without the insertion is first encountered by repair DNA polymerase and is then (mis)used as a template, the palindrome is perfected. When the QP + 4 arm is encountered first, Ura+ derivatives only occur after second-end capture and second-strand synthesis. QP + 4 mutations are suppressed by mismatch repair (MMR) proteins Msh2, Msh3, and Mlh1, but not Msh6. Deleting Rdh54 significantly reduces QP mutations only when events creating Ura+ occur in the context of a D-loop but not during second-strand synthesis. A similar bias is found with a proofreading-defective DNA polymerase mutation (poI3-01). DSB-induced mutations differed in several genetic requirements from spontaneous events. We also created a + 1 frameshift in the donor, expanding a run of 4 Cs to 5 Cs. Again, Ura+ recombinants markedly increased by disabling MMR, suggesting that MMR acts during GC but favors the unbroken, template strand.
摘要：
尽管酿酒酵母中的基因转换（GC）是修复双链断裂（DSB）的最无错误的方法，同源重组过程中的突变率是复制过程中的1000倍。许多突变涉及将部分复制的链从其修复模板上解离，并与相同或另一个模板重新对齐。导致同核苷酸运行中的-1移码，准回文（QP）相关突变和微同源性介导的染色体间模板转换。我们研究了HO内切核酸酶在MATα裂解诱导的GC，由HMR::KI-URA3捐赠者修复。我们在HMR::KI-URA3中插入了一个18bp的反向重复序列，其中一个臂具有4bp的插入。大多数GC产生MAT::KI-ura3::QP+4(Ura-)结果，但是模板切换会产生Ura+菌落，失去了4-bp的插入。如果没有插入的QP臂首先遇到修复DNA聚合酶，然后（错误）用作模板，回文是完善的。当首先遇到QP+4臂时，Ura+衍生物仅在第二末端捕获和第二链合成之后发生。QP+4突变被错配修复(MMR)蛋白Msh2、Msh3和Mlh1抑制,但不被Msh6抑制。仅当创建Ura+的事件发生在D-环的背景下而不是在第二链合成期间发生时,删除Rdh54才显著减少QP突变。在校对缺陷的DNA聚合酶突变（poI3-01）中发现了类似的偏差。DSB诱导的突变在几个遗传要求上与自发事件不同。我们还在捐赠者中创建了+1移码，将4个Cs扩展到5个Cs。再一次,Ura+重组体通过禁用MMR而显著增加，这表明MMR在GC期间起作用，但有利于不间断的，模板链。
