Molecular basis of VCP-linked degeneration

含有瓦洛辛的蛋白（VCP）是几种退行性疾病的致病基因，包括 额颞叶痴呆和ALS，但VCP疾病的分子机制尚不清楚。到 为了研究VCP介导的细胞变性的潜在机制，我们使用CRISPR敲入 基因组工程，以产生果蝇VCP疾病突变的内源性基因置换。 VCP疾病的主要标志是细胞中毒性蛋白聚集体的异常积累。 细胞质，暗示蛋白质清除机制的缺陷。最近，我们和其他实验室发现， 几种VCP疾病突变与自噬介导的降解缺陷有关。我们现在 研究VCP疾病突变如何破坏自噬依赖的蛋白质清除。在我们最初的研究中， 我们有了一个惊人的发现，溶酶体并不单纯以囊泡结构存在， 典型地观察到，但在多种细胞类型中形成膨胀的管状结构，包括肌肉和神经胶质。 这些独特的溶酶体细胞器需要VCP用于其形成和/或维持，因为VCP的过度表达导致VCP的过度表达。 几种致病的VCP突变破坏了溶酶体的管状结构， 破坏自噬体-溶酶体融合。我们假设溶酶体小管的破坏是直接原因 在VCP相关疾病中观察到的自噬缺陷。这项建议的主要目标是：（1） 为了鉴定VCP支持正常溶酶体结构和功能的分子机制， 健康的细胞;（2）阐明这些功能在疾病状态下是如何被破坏的。

Valosin containing protein (VCP) is the causative gene for several degenerative diseases, including frontotemporal dementia and ALS, but the molecular mechanisms underlying VCP diseases are unknown. To investigate potential mechanisms of VCP-mediated cellular degeneration, we are using CRISPR knock-in genome engineering to generate endogenous gene replacements of VCP disease mutations in Drosophila. The major hallmark of VCP diseases is the abnormal accumulation of toxic protein aggregates in the cytoplasm, hinting at a defect in protein clearance mechanisms. Recently, we and other labs have found that several VCP disease mutations are linked to defects in autophagy-mediated degradation. We are now investigating how VCP disease mutations disrupt autophagy-dependent protein clearance. In our initial studies, we made a surprising discovery that lysosomes do not purely exist as vesicular structures, as they are classically viewed, but form expansive tubular structures in multiple cell types, including muscles and glia. These unique lysosomal organelles require VCP for their formation and/or maintenance, as over-expression of several disease-causing VCP mutations collapse the tubular architecture of lysosomes and consequently disrupt autophagosome-lysosome fusion. We hypothesize that disruption of lysosome tubules is a direct cause of the autophagy defects observed in VCP-associated diseases. The broad objectives of this proposal are: (1) to identify the molecular mechanisms by which VCP supports lysosome structure and function in normal, healthy cells; and (2) to clarify how these functions are disrupted in disease states.