Identifying the molecular mechanisms of GEMIN5 mutations in a novel cerebellar ataxia syndrome

摘要 GEMIN5是一种RNA结合蛋白，对组装存活运动神经元(SMN)至关重要 很复杂。GEMIN5促进小核核糖核蛋白的形成 剪接体的构建块)。我们发现了新的常染色体隐性变异体 以运动功能障碍、共济失调和小脑为主要表现的多发性患者的GEMIN5基因 萎缩。 我们提出的研究旨在了解突变的GEMIN5的分子机制 造成我们病人神经异常的原因。我们发现那个病人是个 IPSC患者GEMIN5蛋白水平显著降低和蛋白稳定性降低 神经元提示可能丧失了功能机制。我们的体外组装试验 表明GEMIN5变异体扰乱了SnRNP组装的形成。要了解 功能丧失的后果GEMIN5，我们击倒内源性僵直的死亡， 果蝇与人类GEMIN5的同源物，在果蝇中。击倒僵直的身体(钻机) 导致运动功能障碍、寿命缩短和发育迟缓。有趣的是，我们 观察到辅酶Q10水平在人类患者细胞和我们的 果蝇模型。辅酶Q10治疗可缩短人类GEMIN5的病程 病人。我们使用CRISPR/Cas9建立了Gin5的小鼠模型，并发现早期致死性 在老鼠身上。我们提出的研究旨在了解GEMIN5的分子机制 通过1)对突变的GEMIN5患者神经元进行功能分析；2)检查是否 GEMIN5变异导致体内线粒体功能障碍和IPSC神经元；3) 在小鼠模型中研究GEMIN5突变的机制。我们预计将确定 在人类患者中受到干扰的分子通路。

Abstract GEMIN5, an RNA-binding protein, is essential for assembly of the Survival Motor Neuron (SMN) complex. GEMIN5 facilitates the formation of small nuclear ribonucleoproteins (snRNPs; the building blocks of spliceosomes). We identified novel autosomal recessive variants in the GEMIN5 gene in multiple patients presenting with motor dysfunction, ataxia, and cerebellar atrophy. Our proposed studies are aimed to understand the molecular mechanisms of mutant GEMIN5 responsible for causing the neurological abnormalities in our patients. We found that patient a significant decrease in GEMIN5 protein levels and reduced protein stability in patient iPSC neurons suggesting a possible loss of function mechanism. Our in vitro assembly assay showed that GEMIN5 variants perturb snRNP assembly formation. To understand the consequences of loss of function GEMIN5, we knockdown endogenous rigor mortis, the Drosophila homologue of human GEMIN5, in Drosophila. Knockdown of rigor mortis (rig) caused motor dysfunction, reduced life span and developmental delay. Interestingly, we observed that CoQ10 levels were significantly reduced in human patient cells and our drosophila model. Treatment with CoQ10 reduced the disease course in human GEMIN5 patients. We generated a mouse model of Gemin5 using CRISPR/cas9 and found early lethality in mice. Our proposed studies are aimed to understand the molecular mechanisms of GEMIN5 by 1) conducting functional analysis of mutant GEMIN5 patient neurons; 2) examining if GEMIN5 variants cause mitochondrial dysfunctions in vivo and iPSC neurons; and 3) investigating the mechanisms of GEMIN5 mutations in mouse models. We expect to identify the molecular pathways that are perturbed in human patients.