Ubiquitin-activating enzyme UBA1 is required for cellular response to DNA damage
Ubiquitin-activating enzyme UBA1 is required for cellular response to DNA damage
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DOI：
10.4161/cc.19978
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发表时间：
2012-04-15
期刊：
影响因子：
4.3
通讯作者：
Bartek, Jiri
中科院分区：
文献类型：
--
作者：
Moudry, Pavel;Lukas, Claudia;Bartek, Jiri
细胞 DNA 损伤反应 (DDR) 机制可维持基因组完整性并预防包括癌症在内的严重病变，它是通过蛋白质修饰的信号传导来协调的。蛋白质泛素化调节 DNA 双链断裂 (DSB) 的修复、各种代谢引起的毒性损伤以及电离辐射 (IR) 等环境损害。虽然 DSB 诱发的泛素化级联的几个组成部分已被鉴定，包括 RNF168 和 BRCA1 泛素连接酶，其遗传缺陷分别容易导致类似于共济失调性血管扩张和癌症的综合征，但参与 DDR 的顶端 E1 酶的身份尚未确定。在这里，我们将泛素激活酶 UBA1 确定为人类细胞中响应 IR 和复制应激所需的 E1 酶。我们发现，siRNA 介导的 UBA1 敲低（但不是另一个 UBA 家族成员 UBA6）的敲低，通过 DSB 反应途径的下游组件 53BP1 和 BRCA1 损害了 DNA 损伤和 IR 诱导灶 (IRIF) 位点泛素缀合物的形成。此外，UBA1 的化学抑制可阻止 IRIF 的形成，并严重损害 DSB 修复和 G(1) 中 53BP1 体的形成，G(1) 是复制应激反应的标志。相比之下，DSB 反应的上游步骤，例如组蛋白 H2AX 的磷酸化和 MDC1 的募集，仍然不受 UBA1 耗尽的影响。总体而言，我们的数据表明 UBA1 是对 DSB 泛素化依赖性信号传导和人类细胞复制应激至关重要的顶端酶，对维持基因组完整性、疾病发病机制和癌症治疗具有重要意义。
The cellular DNA damage response (DDR) machinery that maintains genomic integrity and prevents severe pathologies, including cancer, is orchestrated by signaling through protein modifications. Protein ubiquitylation regulates repair of DNA double-strand breaks (DSBs), toxic lesions caused by various metabolic as well as environmental insults such as ionizing radiation (IR). Whereas several components of the DSB-evoked ubiquitylation cascade have been identified, including RNF168 and BRCA1 ubiquitin ligases, whose genetic defects predispose to a syndrome mimicking ataxiatelangiectasia and cancer, respectively, the identity of the apical E1 enzyme involved in DDR has not been established. Here, we identify ubiquitin-activating enzyme UBA1 as the E1 enzyme required for responses to IR and replication stress in human cells. We show that siRNA-mediated knockdown of UBA1, but not of another UBA family member UBA6, impaired formation of both ubiquitin conjugates at the sites of DNA damage and IR-induced foci (IRIF) by the downstream components of the DSB response pathway, 53BP1 and BRCA1. Furthermore, chemical inhibition of UBA1 prevented IRIF formation and severely impaired DSB repair and formation of 53BP1 bodies in G(1), a marker of response to replication stress. In contrast, the upstream steps of DSB response, such as phosphorylation of histone H2AX and recruitment of MDC1, remained unaffected by UBA1 depletion. Overall, our data establish UBA1 as the apical enzyme critical for ubiquitylation-dependent signaling of both DSBs and replication stress in human cells, with implications for maintenance of genomic integrity, disease pathogenesis and cancer treatment.