Replication-Coupled DNA-Protein Crosslink Repair by SPRTN and the Proteasome in Xenopus Egg Extracts
Replication-Coupled DNA-Protein Crosslink Repair by SPRTN and the Proteasome in Xenopus Egg Extracts
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DOI：
10.1016/j.molcel.2018.11.024
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发表时间：
2019-02-07
期刊：
影响因子：
16
通讯作者：
Duxin, Julien P.
中科院分区：
文献类型：
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作者：
Larsen, Nicolai B.;Gao, Alan O.;Duxin, Julien P.
DNA-蛋白质交联(DPC)是干扰DNA新陈代谢的巨大损伤，因此威胁到基因组的完整性。最近的研究表明，金属蛋白酶SPRTN参与了DPC的S时相去除，但在DNA复制过程中SPRTN是如何靶向DPC的，目前还不清楚。利用非洲爪蛙卵提取物概括复制偶联的DPC蛋白分解，我们发现DPC可以被SPRTN或蛋白酶体降解，它们作为独立的DPC蛋白水解酶。蛋白酶体的募集需要DPC的多泛素化，这部分依赖于TRAIP的泛素连接酶活性。相反，SPRTN介导的DPC降解不需要DPC的多泛素化，而是依赖于新生的链延伸到病变的几个核苷酸内，这意味着DPC上的聚合酶停滞激活了领先和滞后链模板上的SPRTN。我们的结果表明，SPRTN和蛋白酶体的活性与DNA复制是通过不同的机制耦合的，这种机制促进了复制跨越不可移动的蛋白质屏障。
DNA-protein crosslinks (DPCs) are bulky lesions that interfere with DNA metabolism and therefore threaten genomic integrity. Recent studies implicate the metalloprotease SPRTN in S phase removal of DPCs, but how SPRTN is targeted to DPCs during DNA replication is unknown. Using Xenopus egg extracts that recapitulate replication-coupled DPC proteolysis, we show that DPCs can be degraded by SPRTN or the proteasome, which act as independent DPC proteases. Proteasome recruitment requires DPC polyubiquitylation, which is partially dependent on the ubiquitin ligase activity of TRAIP. In contrast, SPRTN-mediated DPC degradation does not require DPC polyubiquitylation but instead depends on nascent strand extension to within a few nucleotides of the lesion, implying that polymerase stalling at the DPC activates SPRTN on both leading and lagging strand templates. Our results demonstrate that SPRTN and proteasome activities are coupled to DNA replication by distinct mechanisms that promote replication across immovable protein barriers.