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

鉴定新型小脑共济失调综合征中 GEMIN5 突变的分子机制

基本信息

批准号：
10753403
负责人：
Udai B Pandey
金额：
$ 53.97万
依托单位：
UNIVERSITY OF PITTSBURGH AT PITTSBURGH
依托单位国家：
美国
项目类别：
财政年份：
2023
资助国家：
美国
起止时间：
2023-09-15 至 2028-06-30
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10753403
关键词：
Affect Age of Onset Alternative Splicing Animals Applications Grants Ataxia Atrophic Behavior Biochemical Biogenesis Biological Biological Assay Biological Models Brain CRISPR/Cas technology Cells Cerebellar Ataxia Clinical Coenzyme Q10 Complex DNA Defect Developmental Delay Disorders Disease Drosophila genus Family Fibroblasts Foundations GEMIN5 gene Genetic Guide RNA Heterozygote Homologous Gene Human In Vitro Individual Induced pluripotent stem cell derived neurons Infant Intervention Lead Light Longevity Mitochondria Modeling Molecular Morphology Motor Motor Neuron Disease Motor Neurons Mus Mutation Neurologic Neurons Pathogenesis Pathogenicity Pathway interactions Patients Physiological Process Proteins Proteomics RNA Splicing RNA-Binding Proteins Reporting Respiration Rigor Mortis Role SMN protein (spinal muscular atrophy)SMN1 gene Small Nuclear Ribonucleoproteins Spinal Muscular Atrophy Spliceosomes Symptoms Syndrome Testing Time Transcript Transgenic Mice Transgenic Model Translations Variant autosome clinically relevant disease-causing mutation fly genetic variant human disease in vivo knock-down loss of function mRNA Precursor mitochondrial dysfunction motor disorder motor neuron degeneration mouse model mutant novel pharmacologic polysome profiling premature protein complex protein expression protein function protein protein interaction snRNP Structural Core Protein transcriptome sequencing translatome

项目摘要

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.

抽象的 GEMIN5 是一种 RNA 结合蛋白，对于存活运动神经元 (SMN) 的组装至关重要复杂的。 GEMIN5 促进小核核糖核蛋白（snRNP；剪接体的构建模块）。我们发现了新的常染色体隐性变异多例出现运动功能障碍、共济失调和小脑功能障碍的患者中的 GEMIN5 基因萎缩。我们提出的研究旨在了解突变型 GEMIN5 的分子机制造成我们患者神经系统异常的原因。我们发现该患者患者 iPSC 中 GEMIN5 蛋白水平显着降低且蛋白稳定性降低表明可能存在功能丧失机制的神经元。我们的体外组装试验研究表明 GEMIN5 变异会干扰 snRNP 组装的形成。要了解 GEMIN5 功能丧失的后果，我们抑制内源性尸僵，果蝇中人类 GEMIN5 的果蝇同源物。击倒尸僵（rig）导致运动功能障碍、寿命缩短和发育迟缓。有趣的是，我们观察到人类患者细胞中的 CoQ10 水平显着降低，我们的果蝇模型。 CoQ10 治疗可缩短人类 GEMIN5 的病程患者。我们使用 CRISPR/cas9 生成了 Gemin5 小鼠模型并发现了早期致死性在小鼠中。我们提出的研究旨在了解 GEMIN5 的分子机制 1) 对突变型 GEMIN5 患者神经元进行功能分析； 2）检查是否 GEMIN5变异导致体内和iPSC神经元线粒体功能障碍；和 3) 研究小鼠模型中 GEMIN5 突变的机制。我们期望确定人类患者中受到干扰的分子途径。