The role of the ubiquitin-proteasome system in photoreceptor degeneration

泛素-蛋白酶体系统在光感受器变性中的作用

• 批准号: 9542827
• 负责人: Paige Merritt Dexter
• 金额: $ 3.92万
• 依托单位: DUKE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-09-01 至 2019-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9542827
• 关键词: Affect; Apoptotic; Blindness; Cell Death; Cessation of life; Complex; Data; Development; Disease; Disease Progression; Electroretinography; Equilibrium; Eye; Eye diseases; Functional disorder; Genes; Goals; Immunohistochemistry; Impairment; Individual; Ligase; Link; Mass Spectrum Analysis; Measures; Mediating; Membrane; Methodology; Modeling; Molecular; Molecular Chaperones; Morphology; Mus; Mutant Strains Mice; Mutation; Pathology; Pharmacology; Photoreceptors; Process; Production; Proteasome Inhibitor; Protein Biosynthesis; Proteins; Reporter; Retinal; Retinal Degeneration; Retinitis Pigmentosa; Role; Stress; System; Therapeutic; Therapeutic Intervention; Transgenic Mice; Ubiquitin; Ubiquitination; Western Blotting; Work; effective therapy; experimental study; gene therapy; improved; infancy; inherited retinal degeneration; inhibitor/antagonist; misfolded protein; mouse model; multicatalytic endopeptidase complex; mutant; new therapeutic target; overexpression; photoreceptor degeneration; prevent; protein degradation; proteostasis; retinal rods; targeted treatment; therapeutic target; ubiquitin ligase; ubiquitin-protein ligase

PROJECT SUMMARY Inherited retinal degenerations, including retinitis pigmentosa (RP), originate from mutations in nearly 200 genes affecting photoreceptor cells (rods in the case of RP). Therapeutic options for treating these diseases are limited. One possible approach to treat these conditions is to study the common pathobiological mechanisms downstream of multiple mutations. For example, many mutations linked to retinal degenerations cause photoreceptor proteins to fold poorly and to be targeted to the proteasome for degradation. This suggests a connection between altered cellular proteostasis (i.e. the balance between protein synthesis and degradation) and photoreceptor degeneration. So far, any mechanistic understanding of this connection is still in its infancy. The experiments in this proposal will evaluate the function of the ubiquitin-proteasome system (UPS), the cellular machinery that selectively degrades proteins, in the context of two mouse models of retinal degeneration linked to production of misfolded proteins. Protein degradation by the UPS is typically initiated when a protein is marked for degradation by ubiquitination, which enables the proteasome to recognize and degrade the protein. Many proteins also require processing by molecular complexes involving the chaperone P97 before they can be recognized by proteasomes. Recent work in multiple mouse models of retinal degeneration demonstrated that mutant rods suffer from insufficient UPS capacity prior to the onset of cell death. However, the specific UPS component that is limiting protein degradation in these models remains unknown. The goal of this proposal is to identify this limiting component, which will pave the way for the development of pharmacologic and gene therapy approaches for treatment of inherited retinal degenerations. Aim 1 of this proposal seeks to determine whether UPS-mediated protein degradation in mutant rods is limited by P97. This will be accomplished by measuring the accumulation in mutant rods of a P97-independent reporter of UPS function. Aim 2 will investigate the possibility that the UPS is limited is at the level of ubiquitination by comparing the rate of UbG76VGFP ubiquitination between WT and mutant mice treated with P97 and proteasome inhibitor. In Aim 3 the identified limiting UPS component will be overexpressed in mutant rods, and resulting UPS activity, rod degeneration, and rod function will be evaluated. The results of these Aims will identify the critical component of the UPS whose capacity is overwhelmed prior to rod death. The proposed work represents the first systematic examination of the UPS in the context of ongoing photoreceptor degeneration and will advance the understanding of the factors underlying the pathology of this complex eye disease.

项目摘要 遗传性视网膜变性，包括视网膜色素变性（RP），起源于近200个基因的突变。 影响感光细胞的基因（RP情况下的视杆细胞）。治疗这些疾病的治疗选择是 有限公司治疗这些疾病的一种可能的方法是研究共同的病理生物学机制 多个突变的下游。例如，许多与视网膜变性有关的突变导致 光感受器蛋白折叠不良，并被靶向蛋白酶体降解。这表明 改变细胞蛋白质稳态之间的联系（即蛋白质合成和降解之间的平衡） 和光感受器退化。到目前为止，对这种联系的任何机械理解都还处于初级阶段。 本实验将评估泛素-蛋白酶体系统（UPS）的功能， 选择性降解蛋白质的细胞机制，在两个视网膜病变小鼠模型的背景下， 与产生错误折叠的蛋白质有关的变性。UPS的蛋白质降解通常是在 当蛋白质被标记为通过泛素化降解时，这使得蛋白酶体能够识别并 降解蛋白质。许多蛋白质也需要分子伴侣参与的分子复合物的加工 在它们被蛋白酶体识别之前。视网膜神经病变多个小鼠模型的最新研究 变性表明突变体杆遭受不足的UPS能力之前，细胞的发病 死亡然而，在这些模型中限制蛋白质降解的特定UPS组分仍然存在， 未知本提案的目标是确定这一限制因素，这将为 开发用于治疗遗传性视网膜变性的药理学和基因治疗方法。 本提案的目的1旨在确定突变体视杆细胞中UPS介导的蛋白质降解是否 P97的限制这将通过测量突变体视杆中P97-非依赖性蛋白的积累来实现。 UPS功能报告。目标2将调查UPS受限于以下水平的可能性： 通过比较WT和突变小鼠之间UbG 76 VGFP泛素化的速率， P97和蛋白酶体抑制剂。在目标3中，所鉴定的限制性UPS组分将在以下细胞中过表达： 将评价突变的视杆细胞和产生的UPS活性、视杆细胞变性和视杆细胞功能。的结果 这些目标将确定UPS的关键部件，其容量在棒死亡之前被压倒。 拟议的工作是第一次有系统地审查UPS的背景下， 光感受器变性，并将推进对这种病理学基础因素的理解， 复杂的眼疾