Laboratory of Molecular Pharmacology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892-4255, USA.
EMBO J. 2009 Dec 2;28(23):3667-80. doi: 10.1038/emboj.2009.302. Epub 2009 Oct 22.
Human tyrosyl-DNA phosphodiesterase (TDP1) hydrolyzes the phosphodiester bond at a DNA 3' end linked to a tyrosyl moiety. This type of linkage is found at stalled topoisomerase I (Top1)-DNA covalent complexes, and TDP1 has been implicated in the repair of such complexes. Here we show that Top1-associated DNA double-stranded breaks (DSBs) induce the phosphorylation of TDP1 at S81. This phosphorylation is mediated by the protein kinases: ataxia-telangiectasia-mutated (ATM) and DNA-dependent protein kinase (DNA-PK). Phosphorylated TDP1 forms nuclear foci that co-localize with those of phosphorylated histone H2AX (gammaH2AX). Both Top1-induced replication- and transcription-mediated DNA damages induce TDP1 phosphorylation. Furthermore, we show that S81 phosphorylation stabilizes TDP1, induces the formation of XRCC1 (X-ray cross-complementing group 1)-TDP1 complexes and enhances the mobilization of TDP1 to DNA damage sites. Finally, we provide evidence that TDP1-S81 phosphorylation promotes cell survival and DNA repair in response to CPT-induced DSBs. Together; our findings provide a new mechanism for TDP1 post-translational regulation by ATM and DNA-PK.
人酪氨酸-DNA 磷酸二酯酶（TDP1）可水解与酪氨酸部分相连的 DNA 3'端的磷酸二酯键。这种连接方式存在于拓扑异构酶 I（Top1）-DNA 共价复合物中，并且 TDP1 已被牵连到这些复合物的修复中。在这里，我们发现 Top1 相关的 DNA 双链断裂（DSB）诱导 TDP1 在 S81 处发生磷酸化。这种磷酸化由蛋白激酶：共济失调毛细血管扩张症突变（ATM）和 DNA 依赖性蛋白激酶（DNA-PK）介导。磷酸化的 TDP1 形成核焦点，与磷酸化组蛋白 H2AX（γH2AX）的焦点共定位。Top1 诱导的复制和转录介导的 DNA 损伤均诱导 TDP1 磷酸化。此外，我们还表明，S81 磷酸化稳定 TDP1，诱导 XRCC1（X 射线交叉互补组 1）-TDP1 复合物的形成，并增强 TDP1 向 DNA 损伤部位的募集。最后，我们提供了证据表明，TDP1-S81 磷酸化促进了 CPT 诱导的 DSB 后细胞存活和 DNA 修复。总之，我们的研究结果为 ATM 和 DNA-PK 对 TDP1 的翻译后调节提供了新的机制。
