The helicase domain of Polθ counteracts RPA to promote alt-NHEJ.
The helicase domain of Polθ counteracts RPA to promote alt-NHEJ.
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DOI：
10.1038/nsmb.3494
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发表时间：
2017-12
影响因子：
16.8
通讯作者：
Sfeir A
中科院分区：
文献类型：
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作者：
Mateos-Gomez PA;Kent T;Deng SK;McDevitt S;Kashkina E;Hoang TM;Pomerantz RT;Sfeir A
哺乳动物聚合酶Theta(POL-θ)是一种多功能的酶，通过交替-NHEJ(ALT-NHEJ)促进错误的DNA修复。在这里，我们进行结构-功能分析，并报告，除了聚合酶结构域，解旋酶活性在POLθ介导的双链断裂修复中发挥核心作用。我们的结果表明，POLθ解旋酶通过ALT-NHEJ促进小鼠胚胎干细胞中的染色体易位。解旋酶活性还通过同源重组(HR)抑制CRISPR/Cas9介导的基因靶向。体外实验表明，POLθ-Helicase促进从切除的DSB中去除RPA，以使它们能够退火并随后被ALT-NHEJ连接。与RPA在ALT-NHEJ期间的拮抗作用一致，我们表明RPA1的抑制增强了末端连接并抑制了重组。综上所述，我们的结果揭示了HR和ALT-NHEJ之间的平衡是由Polθ和Rpa相反的活性控制的，这为进一步了解哺乳动物细胞修复途径选择的调节提供了进一步的见解。
Mammalian polymerase theta (Polθ) is a multifunctional enzyme that promotes error-prone DNA repair by alternative-NHEJ (alt-NHEJ). Here we perform structure-function analyses and report that, in addition to the polymerase domain, the helicase activity plays a central role during Polθ-mediated double-strand break (DSB) repair. Our results show that Polθ-helicase promotes chromosomal translocations by alt-NHEJ in mouse embryonic stem cells. The helicase activity also suppresses CRISPR/Cas9 mediated gene targeting by homologous recombination (HR). In vitro experiments reveal that Polθ–helicase facilitates the removal of RPA from resected DSBs to allow their annealing and subsequent joining by alt-NHEJ. Consistent with an antagonistic role for RPA during alt-NHEJ, we show that the inhibition of RPA1 enhances end-joining and suppresses recombination. Taken together, our results reveal that the balance between HR and alt-NHEJ is controlled by opposing activities of Polθ and RPA, providing further insight into the regulation of repair pathway choice in mammalian cells.
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影响因子：
64.8
作者：
Mateos-Gomez PA;Gong F;Nair N;Miller KM;Lazzerini-Denchi E;Sfeir A
通讯作者：
Sfeir A
影响因子：
16.8
作者：
Kent, Tatiana;Chandramouly, Gurushankar;McDevitt, Shane Michael;Ozdemir, Ahmet Y.;Pomerantz, Richard T.
通讯作者：
Pomerantz, Richard T.
影响因子：
16.6
作者：
Bothmer A;Phadke T;Barrera LA;Margulies CM;Lee CS;Buquicchio F;Moss S;Abdulkerim HS;Selleck W;Jayaram H;Myer VE;Cotta-Ramusino C
通讯作者：
Cotta-Ramusino C
影响因子：
4.5
作者：
Chan SH;Yu AM;McVey M
通讯作者：
McVey M
影响因子：
64.8
作者：
通讯作者：
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