von Stechow Louise, Typas Dimitris, Carreras Puigvert Jordi, Oort Laurens, Siddappa Ramakrishnaiah, Pines Alex, Vrieling Harry, van de Water Bob, Mullenders Leon H F, Danen Erik H J
Division of Toxicology, Leiden/Amsterdam Center for Drug Research, Leiden University, Leiden, The Netherlands.
Department of Toxicogenetics, Leiden University Medical Center, Leiden, The Netherlands.
Mol Cell Biol. 2015 Apr;35(7):1254-68. doi: 10.1128/MCB.01152-14. Epub 2015 Jan 26.
DNA damage response signaling is crucial for genome maintenance in all organisms and is corrupted in cancer. In an RNA interference (RNAi) screen for (de)ubiquitinases and sumoylases modulating the apoptotic response of embryonic stem (ES) cells to DNA damage, we identified the E3 ubiquitin ligase/ISGylase, ariadne homologue 1 (ARIH1). Silencing ARIH1 sensitized ES and cancer cells to genotoxic compounds and ionizing radiation, irrespective of their p53 or caspase-3 status. Expression of wild-type but not ubiquitinase-defective ARIH1 constructs prevented sensitization caused by ARIH1 knockdown. ARIH1 protein abundance increased after DNA damage through attenuation of proteasomal degradation that required ATM signaling. Accumulated ARIH1 associated with 4EHP, and in turn, this competitive inhibitor of the eukaryotic translation initiation factor 4E (eIF4E) underwent increased nondegradative ubiquitination upon DNA damage. Genotoxic stress led to an enrichment of ARIH1 in perinuclear, ribosome-containing regions and triggered 4EHP association with the mRNA 5' cap as well as mRNA translation arrest in an ARIH1-dependent manner. Finally, restoration of DNA damage-induced translation arrest in ARIH1-depleted cells by means of an eIF2 inhibitor was sufficient to reinstate resistance to genotoxic stress. These findings identify ARIH1 as a potent mediator of DNA damage-induced translation arrest that protects stem and cancer cells against genotoxic stress.
DNA损伤反应信号传导对于所有生物体的基因组维持至关重要，而在癌症中则会受损。在一项针对调节胚胎干细胞（ES）对DNA损伤的凋亡反应的（去）泛素酶和SUMO化酶的RNA干扰（RNAi）筛选中，我们鉴定出E3泛素连接酶/ISGylase，即阿里阿德涅同源物1（ARIH1）。沉默ARIH1会使ES细胞和癌细胞对基因毒性化合物和电离辐射敏感，无论它们的p53或caspase-3状态如何。野生型而非泛素酶缺陷型ARIH1构建体的表达可防止由ARIH1敲低引起的敏感性。DNA损伤后，ARIH1蛋白丰度通过需要ATM信号传导的蛋白酶体降解减弱而增加。积累的ARIH1与4EHP相关，反过来，这种真核翻译起始因子4E（eIF4E）的竞争性抑制剂在DNA损伤时经历了增加的非降解性泛素化。基因毒性应激导致ARIH1在核周含核糖体区域富集，并以ARIH1依赖的方式触发4EHP与mRNA 5'帽的结合以及mRNA翻译停滞。最后，通过eIF2抑制剂恢复ARIH1缺失细胞中DNA损伤诱导的翻译停滞足以恢复对基因毒性应激的抗性。这些发现确定ARIH1是DNA损伤诱导的翻译停滞的有效介质，可保护干细胞和癌细胞免受基因毒性应激。
