Genome-wide analysis of Rad52 foci reveals diverse mechanisms impacting recombination.
Genome-wide analysis of Rad52 foci reveals diverse mechanisms impacting recombination.
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DOI：
10.1371/journal.pgen.0030228
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发表时间：
2007-12
期刊：
影响因子：
4.5
通讯作者：
Rothstein R
中科院分区：
文献类型：
--
作者：
Alvaro D;Lisby M;Rothstein R
为了研究DNA损伤反应，我们在酿酒酵母中进行了全基因组研究，并确定了86个基因缺失，导致增殖二倍体细胞中自发Rad 52灶水平增加。从酵母到人类，超过一半的基因是保守的。沿着与DNA复制、修复和染色质重塑相关的基因，我们发现了22个以前未表征的开放阅读框架。重组率和与rad 52 Δ的合成遗传相互作用的分析表明，多种机制是导致自发Rad 52灶水平升高的原因，包括重组发生性病变的产生增加、姐妹染色单体重组缺陷和不正确的灶组装/拆卸。我们的细胞生物学方法证明了同源重组过程的多样性，保护免受自发DNA损伤，并促进有效的修复。同源重组（HR）是一个细胞过程，允许有效修复内源性和外源性DNA损伤。虽然人力资源的主要参与者已经很好地描述了，但不同过程与人力资源途径的相互作用仍然是神秘的。传统上，研究HR的遗传筛选利用遗传测定，例如暴露于DNA损伤剂后的存活率或重组产物产生速率的改变。在这项工作中，我们而是利用细胞生物学表型，中央HR蛋白Rad 52的重新定位到亚核灶反映修复中心积极从事HR. This方法使我们能够识别突变体，影响动力学的HR修复中心的组装和拆卸，无论重组的结果。我们鉴定了86个导致增殖二倍体细胞中自发灶水平增加的基因缺失，其中22个是先前未表征的ORF的缺失（命名为IRC 2 -11，13-16，18-25）。突变体的遗传特性揭示了焦点表型的基础机制的多样性。这些包括增加DNA损伤的产生，阻断HR的完成，以及改变遗传重组的动力学和HR蛋白复合物的组装/分解。
To investigate the DNA damage response, we undertook a genome-wide study in Saccharomyces cerevisiae and identified 86 gene deletions that lead to increased levels of spontaneous Rad52 foci in proliferating diploid cells. More than half of the genes are conserved across species ranging from yeast to humans. Along with genes involved in DNA replication, repair, and chromatin remodeling, we found 22 previously uncharacterized open reading frames. Analysis of recombination rates and synthetic genetic interactions with rad52Δ suggests that multiple mechanisms are responsible for elevated levels of spontaneous Rad52 foci, including increased production of recombinogenic lesions, sister chromatid recombination defects, and improper focus assembly/disassembly. Our cell biological approach demonstrates the diversity of processes that converge on homologous recombination, protect against spontaneous DNA damage, and facilitate efficient repair. Homologous recombination (HR) is a cellular process that permits efficient repair of both endogenous and exogenous DNA damage. Although the principal players in HR have been well characterized, the interplay of diverse processes with the HR pathway remains mysterious. Traditionally, genetic screens investigating HR have utilized genetic assays, such as survival following exposure to DNA damaging agents or alterations in the rate of the generation of recombinant products. In this work, we instead utilize a cell biology phenotype, the relocalization of the central HR protein Rad52 into subnuclear foci reflecting repair centers actively engaged in HR. This approach allows us to identify mutants that affect the kinetics of HR repair center assembly and disassembly regardless of the outcome of recombination. We identified 86 gene deletions that lead to increases in the levels of spontaneous foci in proliferating diploid cells, 22 of which were deletions of previously uncharacterized ORFs (designated IRC2–11, 13–16, 18–25). Genetic characterization of the mutants revealed a diversity of mechanisms that underlie the focus phenotype. These include increasing the generation of DNA lesions, blocking the completion of HR, and altering the kinetics of genetic recombination and the assembly/disassembly of the HR protein complexes.
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DOI：
10.1126/science.1135862
发表时间：
2007-02-02
期刊：
Science (New York, N.Y.)
影响因子：
--
作者：
Driscoll R;Hudson A;Jackson SP
通讯作者：
Jackson SP
影响因子：
5.3
作者：
GANGLOFF, S;MCDONALD, JP;ROTHSTEIN, R
通讯作者：
ROTHSTEIN, R
影响因子：
64.8
作者：
Collins, Sean R.;Miller, Kyle M.;Krogan, Nevan J.
通讯作者：
Krogan, Nevan J.
影响因子：
5.3
作者：
Bastin-Shanower, SA;Fricke, WM;Brill, SJ
通讯作者：
Brill, SJ
影响因子：
10.5
作者：
Kaliraman, V;Mullen, JR;Brill, SJ
通讯作者：
Brill, SJ