Genome-wide siRNA screening reveals that DCAF4-mediated ubiquitination of optineurin stimulates autophagic degradation of Cu/Zn superoxide dismutase.
Genome-wide siRNA screening reveals that DCAF4-mediated ubiquitination of optineurin stimulates autophagic degradation of Cu/Zn superoxide dismutase.
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全基因组 siRNA 筛选揭示 DCAF4 介导的 optineurin 泛素化刺激 Cu/Zn 超氧化物歧化酶的自噬降解。
DOI：
10.1074/jbc.ra119.010239
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发表时间：
2020
期刊：
影响因子：
--
通讯作者：
H.
中科院分区：
文献类型：
--
作者：
Homma;K.;Takahashi;H.;Tsuburaya;N.;Naguro;I.;Fujisawa;T. and Ichijo;H.
铜锌超氧化物歧化酶（SOD 1）是神经退行性疾病肌萎缩侧索硬化症（ALS）的致病基因之一。虽然SOD 1突变体（SOD 1 mut）诱导的运动神经元毒性的确切机制尚不清楚，但已知SOD 1蛋白稳态缺陷在ALS发病机制中具有关键作用。我们以前报道过，SOD 1 mutadopts的构象，暴露Derlin-1结合区（DBR）和DBR暴露的SOD 1与Derlin-1相互作用，导致运动神经元死亡。我们还发现，环境变化，即锌耗尽，诱导WT SOD 1（SOD 1 WT）的DBR暴露的构象的构象变化，这表明DBR掩蔽和DBR暴露状态之间的平衡状态的存在，即使与SOD 1 WT。在此，我们进行了基于时间分辨FRET的高通量筛选以进一步研究SOD 1 WT的构象变化，并且我们使用全基因组siRNA筛选来寻找SOD 1蛋白质稳定的调节剂。该筛选产生了30个候选基因，其保持了DBR暴露的SOD 1 WT构象的缺失。在这些基因中，有一个编码DDB 1和CUL 4相关因子4（DCAF 4），这是E3泛素-蛋白连接酶复合物的底物受体。值得注意的是，我们发现DCAF 4介导ALS相关蛋白和自噬受体视神经磷酸酶（OPTN）的泛素化，并促进DBR暴露的SOD 1的自噬降解。总之，我们的筛选鉴定出DCAF 4是DBR暴露的SOD 1的适当蛋白质稳定所需的，这可能与管理ALS的疗法的开发具有潜在的相关性。
Cu, Zn superoxide dismutase(SOD1) is one of the genes implicated in the devastating neurodegenerative disorder amyotrophic lateral sclerosis (ALS). Although the precise mechanisms ofSOD1mutant (SOD1mut)-induced motoneuron toxicity are still unclear, defects in SOD1 proteostasis are known to have a critical role in ALS pathogenesis. We previously reported that the SOD1mutadopts a conformation that exposes a Derlin-1–binding region (DBR) and that DBR-exposed SOD1 interacts with Derlin-1, leading to motoneuron death. We also found that an environmental change,i.e.zinc depletion, induces a conformational change in WT SOD1 (SOD1WT) to the DBR-exposed conformation, suggesting the presence of an equilibrium state between the DBR-masked and DBR-exposed states even with SOD1WT. Here, we conducted a high-throughput screening based on time-resolved FRET to further investigate the SOD1WTconformational change, and we used a genome-wide siRNA screen to search for regulators of SOD1 proteostasis. This screen yielded 30 candidate genes that maintained an absence of the DBR-exposed SOD1WTconformation. Among these genes was one encoding DDB1- and CUL4-associated factor 4 (DCAF4), a substrate receptor of the E3 ubiquitin–protein ligase complex. Of note, we found that DCAF4 mediates the ubiquitination of an ALS-associated protein and autophagy receptor, optineurin (OPTN), and facilitates autophagic degradation of DBR-exposed SOD1. In summary, our screen identifies DCAF4 as being required for proper proteostasis of DBR-exposed SOD1, which may have potential relevance for the development of therapies for managing ALS.