REV7 is essential for DNA damage tolerance via two REV3L binding sites in mammalian DNA polymerase ζ.
REV7 is essential for DNA damage tolerance via two REV3L binding sites in mammalian DNA polymerase ζ.
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DOI：
10.1093/nar/gku1385
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发表时间：
2015-01
影响因子：
14.9
通讯作者：
Wood RD
中科院分区：
文献类型：
--
作者：
Tomida J;Takata K;Lange SS;Schibler AC;Yousefzadeh MJ;Bhetawal S;Dent SY;Wood RD
DNA聚合酶zeta（polymerase zeta）对于控制突变和遗传不稳定性是非常重要的。REV 3L包含催化亚基，而REV 7（MAD 2L 2）被认为是辅助亚基。然而，REV 7在DNA损伤耐受中的作用与哺乳动物的聚合酶之间是否存在必然的联系尚不明确，越来越多的证据表明REV 7和REV 3L具有独立的功能。由于REV 3L在哺乳动物细胞中表达困难，以及缺乏功能互补系统，阻碍了聚合酶的分析。在这里，我们报告，REV 7与全长REV 3L在体内相互作用，我们确定了一个新的保守的REV 7相互作用位点在人REV 3L（残基1993-2003），不同于已知的结合位点（残基1877-1887）。两个REV 7结合位点的突变消除了REV 3L-REV 7相互作用。体内互补表明，REV 3L中的两个REV 7结合位点对于防止自发染色体断裂和赋予对UV辐射和顺铂的抗性是必需的。这证明了REV 7在聚合酶中的损伤特异性功能，与REV 3L和REV 7在原代细胞活力和胚胎发生中的不同作用相反。
DNA polymerase zeta (pol ζ) is exceptionally important for controlling mutagenesis and genetic instability. REV3L comprises the catalytic subunit, while REV7 (MAD2L2) is considered an accessory subunit. However, it has not been established that the role of REV7 in DNA damage tolerance is necessarily connected with mammalian pol ζ, and there is accumulating evidence that REV7 and REV3L have independent functions. Analysis of pol ζ has been hampered by difficulties in expression of REV3L in mammalian cells, and lack of a functional complementation system. Here, we report that REV7 interacts with full-length REV3L in vivo and we identify a new conserved REV7 interaction site in human REV3L (residues 1993–2003), distinct from the known binding site (residues 1877–1887). Mutation of both REV7-binding sites eliminates the REV3L–REV7 interaction. In vivo complementation shows that both REV7-binding sites in REV3L are necessary for preventing spontaneous chromosome breaks and conferring resistance to UV radiation and cisplatin. This demonstrates a damage-specific function of REV7 in pol ζ, in contrast to the distinct roles of REV3L and REV7 in primary cell viability and embryogenesis.
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