A <em>Drosophila</em> Model of ALS: Human ALS-Associated Mutation in VAP33A Suggests a Dominant Negative Mechanism

ALS8 is caused by a dominant mutation in an evolutionarily conserved protein, VAPB (vesicle-associated membrane protein (VAMP)-associated membrane protein B)/ALS8). We have established a fly model of ALS8 using the corresponding mutation in Drosophila VAPB (dVAP33A) and examined the effects of this mutation on VAP function using genetic and morphological analyses. By simultaneously assessing the effects of VAPwt and VAPP58S on synaptic morphology and structure, we demonstrate that the phenotypes produced by neuronal expression of VAPP58S resemble VAP loss of function mutants and are opposite those of VAP overexpression, suggesting that VAPP58S may function as a dominant negative. This is brought about by aggregation of VAPP58S and recruitment of wild type VAP into these aggregates. Importantly, we also demonstrate that the ALS8 mutation in dVAP33A interferes with BMP signaling pathways at the neuromuscular junction, identifying a new mechanism underlying pathogenesis of ALS8. Furthermore, we show that mutant dVAP33A can serve as a powerful tool to identify genetic modifiers of VAPB. This new fly model of ALS, with its robust pathological phenotypes, should for the first time allow the power of unbiased screens in Drosophila to be applied to study of motor neuron diseases.肌萎缩侧索硬化8型（ALS8）由进化保守蛋白VAPB（囊泡相关膜蛋白（VAMP）相关膜蛋白B）/ALS8的显性突变所导致。我们利用果蝇VAPB同源蛋白dVAP33A中的对应突变构建了ALS8的果蝇模型，并通过遗传学与形态学分析探究了该突变对VAP功能的影响。通过同时评估野生型VAP（VAPwt）与突变型VAPP58S对突触形态与结构的作用，我们发现神经元表达VAPP58S所产生的表型与VAP功能缺失突变体相似，且与VAP过表达的表型完全相反，这提示VAPP58S可能以显性负效应的方式发挥功能。该效应由VAPP58S的聚集以及野生型VAP被招募至这些聚集体中所介导。重要的是，我们还证实dVAP33A中的ALS8突变会干扰神经肌肉接头处的骨形态发生蛋白（BMP）信号通路，从而明确了ALS8发病机制的全新机制。此外，我们证明突变型dVAP33A可作为高效工具，用于筛选VAPB的遗传修饰因子。这款具有显著病理表型的新型ALS果蝇模型，首次使得果蝇的无偏筛选技术得以应用于运动神经元疾病的研究。