Microarray-based genetic screen defines SAW1, a gene required for Rad1/Rad10-dependent processing of recombination intermediates
Microarray-based genetic screen defines SAW1, a gene required for Rad1/Rad10-dependent processing of recombination intermediates
复制标题
DOI：
10.1016/j.molcel.2008.02.028
复制
发表时间：
2008-05-09
期刊：
影响因子：
16
通讯作者：
Lee, Sang Eun
中科院分区：
文献类型：
--
作者：
Li, Fuyang;Dong, Junchao;Lee, Sang Eun
通过单链退火（SSA）介导两侧为同向重复序列的双链断裂（DSB）的消除，其依赖于涉及重组、错配修复和核苷酸切除修复的一组不同的基因产物。在这里，我们筛选酵母突变体缺陷SSA与质粒为基础的SSA检测耦合到条形码微阵列读数。筛选鉴定了Yal 027 Wp/Saw 1（单链退火弱化1）和Slx 4以及其他已知的SSA蛋白。Saw 1与Rad 1/Rad 10、Msh 2/Msh 3和Rad 52蛋白发生物理相互作用，缺乏SLX 4或SAW 1的细胞积累重组中间体，这些重组中间体在Rad 1/Rad 10依赖性3'瓣切割步骤中被阻断。Slx 4和Saw 1也有助于核糖体DNA阵列的完整性。Saw 1突变体不能与Rad 1相互作用，但保留与Rad 52和Msh 2的相互作用，在3'瓣去除和SSA修复中有缺陷。删除SAW 1废除协会的Rad 1 SSA中间体在体内。我们建议，锯目标Rad 1/Rad 10的Rad 52包被的重组中间体。
Elimination of a double-strand break (DSB) flanked by direct repeat sequences is mediated by single-strand annealing (SSA), which relies on a distinct set of gene products involving recombination, mismatch repair, and nucleotide excision repair. Here, we screened for yeast mutants defective in SSA with a plasmid-based SSA assay coupled to a barcode microarray readout. The screen identified Yal027Wp/Saw1 (single-strand annealing weakened 1) and Slx4 besides other known SSA proteins. Saw1 interacts physically with Rad1/Rad10, Msh2/Msh3, and Rad52 proteins, and cells lacking SLX4 orSAW1 accumulate recombination intermediates blocked at the Rad1/Rad10-dependent 3' flap cleavage step. Slx4 and Saw1 also contribute to the integrity of ribosomal DNA arrays. Saw1 mutants that fail to interact with Rad1, but retain interaction with Rad52 and Msh2, are defective in 3' flap removal and SSA repair. Deletion of SAW1 abolished association of Rad1 at SSA intermediates in vivo. We propose that Saw1 targets Rad1/Rad10 to Rad52-coated recombination intermediates.