C9orf72 protein quality control by UBR5-mediated heterotypic ubiquitin chains

Hexanucleotide repeat expansions within C9orf72 are a frequent cause of amyotrophic lateral sclerosis and frontotemporal dementia. Haploinsufficiency leading to reduced C9orf72 protein contributes to disease pathogenesis. C9orf72 binds SMCR8 to form a robust complex that regulates small GTPases, lysosomal integrity and autophagy. In contrast to this functional understanding, we know far less about assembly and turnover of the C9orf72-SMCR8 complex. Loss of either subunit causes the concurrent ablation of the respective partner. However, the molecular mechanism underlying this interdependence remains elusive. Here, we identify C9orf72 as substrate of branched ubiquitin chain-dependent protein quality control. We find that SMCR8 prevents C9orf72 from rapid degradation by the proteasome. Mass spectrometry and biochemical analyses reveal the E3 ligase UBR5 and the BAG6 chaperone complex as C9orf72 interacting proteins, which are components of the machinery that modifies proteins with K11/K48-linked heterotypic ubiquitin chains. Depletion of UBR5 results in reduced K11/K48 ubiquitination and increased C9orf72 when SMCR8 is absent. Our data provides novel insights into C9orf72 regulation with potential implication for strategies to antagonize C9orf72 loss during disease progression.

C9orf72基因内的六核苷酸重复扩增（hexanucleotide repeat expansions）是肌萎缩侧索硬化（amyotrophic lateral sclerosis）与额颞叶痴呆（frontotemporal dementia）的常见致病原因。单倍剂量不足（haploinsufficiency）导致C9orf72蛋白表达量降低，这一过程参与疾病的发病机制。C9orf72可与SMCR8结合形成稳定复合物，该复合物可调控小GTP酶（small GTPases）、溶酶体完整性（lysosomal integrity）与自噬（autophagy）过程。尽管对该复合物的功能已有上述认知，但学界对C9orf72-SMCR8复合物的组装与蛋白周转过程仍知之甚少。任一亚基的缺失都会导致其对应结合亚基同步被降解，然而这种相互依存关系背后的分子机制仍不明确。本研究证实C9orf72是分支泛素链依赖性蛋白质质量控制（protein quality control）的底物。研究发现，SMCR8可阻止C9orf72被蛋白酶体（proteasome）快速降解。质谱分析（mass spectrometry）与生化实验结果显示，E3泛素连接酶（E3 ligase）UBR5与BAG6伴侣复合物（chaperone complex）为C9orf72的互作蛋白；二者均属于以K11/K48连接的异型泛素链（K11/K48-linked heterotypic ubiquitin chains）修饰蛋白质的调控机器组分。在SMCR8缺失的情况下，敲低UBR5会导致K11/K48位泛素化（ubiquitination）水平降低，同时使C9orf72蛋白水平升高。本研究数据为C9orf72的调控机制提供了新见解，同时为对抗疾病进程中C9orf72表达缺失的治疗策略提供了潜在思路。