Break-induced ATR and Ddb1-Cul4Cdt2 ubiquitin ligase-dependent nucleotide synthesis promotes homologous recombination repair in fission yeast
Break-induced ATR and Ddb1-Cul4Cdt2 ubiquitin ligase-dependent nucleotide synthesis promotes homologous recombination repair in fission yeast
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DOI：
10.1101/gad.1970810
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发表时间：
2010-12-01
影响因子：
10.5
通讯作者：
Humphrey, Timothy C.
中科院分区：
文献类型：
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作者：
Moss, Jennifer;Tinline-Purvis, Helen;Humphrey, Timothy C.
核苷酸合成是对DNA损伤的普遍反应，但这种反应如何促进DNA修复和细胞生存尚不清楚。在这里，我们确定了DNA损伤诱导的核苷酸合成在分裂酵母的同源重组(HR)修复中的作用。通过遗传筛选，我们发现DDB1-CUL4(CDT2)泛素连接酶复合体和核糖核苷酸还原酶(RNR)是DNA双链断裂(DSB)HR修复所必需的。DDB1-CUL4(Cdt2)泛素连接酶复合体是裂殖酵母降解RNR抑制剂Spd1所必需的。因此，删除spd1(+)抑制了DNA损伤敏感性，并抑制了与DDB1(+)或cdt2(+)丢失相关的HR效率降低。此外，我们证明了在HR修复过程中，核苷酸合成在切除的单链DNA末端的突触后间隙填充中所起的作用。最后，我们确定了Rad3(ATR)在核苷酸合成和HR中的作用，它通过增加CDT2核水平来响应DNA损伤。我们的发现支持一种模型，在该模型中，Break诱导了Rad3和DDB1-CUL4(CDT2)泛素连接酶依赖的Spd1降解，RNR激活促进了HR修复过程中突触后单链DNA缺口的填补。
Nucleotide synthesis is a universal response to DNA damage, but how this response facilitates DNA repair and cell survival is unclear. Here we establish a role for DNA damage-induced nucleotide synthesis in homologous recombination (HR) repair in fission yeast. Using a genetic screen, we found the Ddb1-Cul4(Cdt2) ubiquitin ligase complex and ribonucleotide reductase (RNR) to be required for HR repair of a DNA double-strand break (DSB). The Ddb1-Cul4(Cdt2) ubiquitin ligase complex is required for degradation of Spd1, an inhibitor of RNR in fission yeast. Accordingly, deleting spd1(+) suppressed the DNA damage sensitivity and the reduced HR efficiency associated with loss of ddb1(+) or cdt2(+). Furthermore, we demonstrate a role for nucleotide synthesis in postsynaptic gap filling of resected ssDNA ends during HR repair. Finally, we define a role for Rad3 (ATR) in nucleotide synthesis and HR through increasing Cdt2 nuclear levels in response to DNA damage. Our findings support a model in which break-induced Rad3 and Ddb1-Cul4(Cdt2) ubiquitin ligase-dependent Spd1 degradation and RNR activation promotes postsynaptic ssDNA gap filling during HR repair.