Molecular Toxicology Group, Department of Biology, University of Konstanz, Konstanz, Germany.
Sci Rep. 2019 Jul 11;9(1):10075. doi: 10.1038/s41598-019-46358-7.
The prime function of nucleoli is ribogenesis, however, several other, non-canonical functions have recently been identified, including a role in genotoxic stress response. Upon DNA damage, numerous proteins shuttle dynamically between the nucleolus and the nucleoplasm, yet the underlying molecular mechanisms are incompletely understood. Here, we demonstrate that PARP1 and PARylation contribute to genotoxic stress-induced nucleolar-nucleoplasmic shuttling of key genome maintenance factors in HeLa cells. Our work revealed that the RECQ helicase, WRN, translocates from nucleoli to the nucleoplasm upon treatment with the oxidizing agent HO, the alkylating agent 2-chloroethyl ethyl sulfide (CEES), and the topoisomerase inhibitor camptothecin (CPT). We show that after treatment with HO and CEES, but not CPT, WRN translocation was dependent on PARP1 protein, yet independent of its enzymatic activity. In contrast, nucleolar-nucleoplasmic translocation of the base excision repair protein, XRCC1, was dependent on both PARP1 protein and its enzymatic activity. Furthermore, gossypol, which inhibits PARP1 activity by disruption of PARP1-protein interactions, abolishes nucleolar-nucleoplasmic shuttling of WRN, XRCC1 and PARP1, indicating the involvement of further upstream factors. In conclusion, this study highlights a prominent role of PARP1 in the DNA damage-induced nucleolar-nucleoplasmic shuttling of genome maintenance factors in HeLa cells in a toxicant and protein-specific manner.
核仁的主要功能是核糖体的生物合成，但最近又发现了其他几种非典型功能，包括在遗传毒性应激反应中的作用。在 DNA 损伤后，许多蛋白质在核仁与核质之间动态穿梭，但潜在的分子机制尚不完全清楚。在这里，我们证明 PARP1 和 PARylation 参与了 HeLa 细胞中遗传毒性应激诱导的关键基因组维护因子的核仁-核质穿梭。我们的工作表明，RECQ 解旋酶 WRN 在氧化应激剂 HO、烷化剂 2-氯乙基乙基硫醚（CEES）和拓扑异构酶抑制剂喜树碱（CPT）处理后，从核仁转移到核质。我们表明，在 HO 和 CEES 处理后，但不是 CPT 处理后，WRN 易位依赖于 PARP1 蛋白，但不依赖其酶活性。相比之下，碱基切除修复蛋白 XRCC1 的核仁-核质易位既依赖于 PARP1 蛋白，也依赖于其酶活性。此外，抑制 PARP1 蛋白相互作用的 gossypol 可破坏 PARP1 对 WRN、XRCC1 和 PARP1 的核仁-核质易位，表明存在其他上游因素的参与。总之，本研究强调了 PARP1 在 HeLa 细胞中 DNA 损伤诱导的基因组维护因子的核仁-核质穿梭中的重要作用，这种作用具有毒物和蛋白特异性。
