Targeting nuclear factor-kappa B (NF-κB) represents a highly viable strategy against chemoresistance in cancers as well as cell death. Ubiquitination, including linear ubiquitination mediated by the linear ubiquitin chain assembly complex (LUBAC), is emerging as a crucial mechanism of overactivated NF-κB signaling. Ovarian tumor family deubiquitinase OTULIN is the only linear linkage–specific deubiquitinase; however, the molecular mechanisms of how it counteracts LUBAC-mediated NF-κB activation have been largely unknown. Here, we identify Lys64/66 of OTULIN for linear ubiquitination facilitated in a LUBAC-dependent manner as a necessary event required for OTULIN–LUBAC interaction under unstressed conditions, which becomes deubiquitinated by OTULIN itself in response to genotoxic stress. Furthermore, this self-deubiquitination of OTULIN occurs intermolecularly, mediated by OTULIN dimerization, resulting in the subsequent dissociation of OTULIN from the LUBAC complex and NF-κB overactivation. Oxidative stress induces OTULIN dimerization via cysteine-mediated covalent disulfide bonds. Our study reveals that the status of the physical interaction between OTULIN and LUBAC is a crucial determining factor for the genotoxic NF-κB signaling, as measured by cell survival and proliferation, while OTULIN loss of function resulting from its dimerization and deubiquitination leads to a dissociation of OTULIN from the LUBAC complex. Of note, similar molecular mechanisms apply to the inflammatory NF-κB signaling in response to tumor necrosis factor α. Hence, a fuller understanding of the detailed molecular mechanisms underlying the disruption of the OTULIN–LUBAC interaction will be instrumental for developing future therapeutic strategies against cancer chemoresistance and necroptotic processes pertinent to numerous human diseases.

中文翻译：

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OTULIN–LUBAC 之间的相互作用决定了基因毒性和炎症性 NF-κB 信号反应

靶向核因子-kappa B (NF-κB) 代表了一种高度可行的策略来对抗癌症的化学抗性和细胞死亡。泛素化，包括由线性泛素链组装复合物 (LUBAC) 介导的线性泛素化，正在成为过度激活 NF-κB 信号传导的重要机制。卵巢肿瘤家族去泛素化酶 OTULIN 是唯一的线性连锁特异性去泛素化酶；然而，它如何抵消 LUBAC 介导的 NF-κB 激活的分子机制在很大程度上是未知的。在这里，我们将 OTULIN 的 Lys64/66 识别为以 LUBAC 依赖性方式促进的线性泛素化，这是在无应激条件下 OTULIN-LUBAC 相互作用所需的必要事件，它被 OTULIN 本身去泛素化以响应基因毒性应激。此外，OTULIN 的这种自去泛素化发生在分子间，由 OTULIN 二聚化介导，导致随后 OTULIN 从 LUBAC 复合物解离和 NF-κB 过度激活。氧化应激通过半胱氨酸介导的共价二硫键诱导 OTULIN 二聚化。我们的研究表明，OTULIN 和 LUBAC 之间物理相互作用的状态是基因毒性 NF-κB 信号传导的关键决定因素，通过细胞存活和增殖来衡量，而 OTULIN 因二聚化和去泛素化而导致的功能丧失导致解离来自 LUBAC 复合物的 OTULIN。值得注意的是，类似的分子机制适用于炎症性 NF-κB 信号转导以响应肿瘤坏死因子 α。因此，