ER stress-induced translational regulation in retinal degeneration

视网膜变性中内质网应激诱导的翻译调节

基本信息

批准号：
10653971
负责人：
Deepika Vasudevan
金额：
$ 23.66万
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：
2021
资助国家：
美国
起止时间：
2021-07-01 至 2024-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10653971
关键词：
5&apos Untranslated Regions Acute Address Affect Age Age related macular degeneration Atherosclerosis Biochemical Blindness Cellular Stress Cessation of life Chronic Complex Cytoprotection Data Destinations Development Diabetes Mellitus Disease Drosophila genus EIF-2alpha Embryo Endoplasmic Reticulum Etiology Feedback Fibroblasts Functional disorder G-Protein-Coupled Receptors Genes Genetic Genetic Diseases Genetic Screening Genetic Transcription Goals Heritability Homeostasis Impairment Integral Membrane Protein Lead Learning Light Mammalian Cell Mediating Membrane Mentors Modeling Molecular Molecular Chaperones Mus Mutation Nature Neurodegenerative Disorders Open Reading Frames Pathologic Pathway interactions Patients Peptide Initiation Factors Pharmacologic Substance Phase Phosphorylation Phosphotransferases Photoreceptors Phylogenetic Analysis Property Protein Synthesis Inhibition Proteins RNA interference screen Regulation Reporter Research Retinal Degeneration Retinitis Pigmentosa Rhodopsin Role Signal Transduction Stress System Techniques Testing Therapeutic Training Transcript Transfer RNA Translating Translational Regulation Translational Repression Translations Work Yeasts age related attenuation autosome biological adaptation to stress design endoplasmic reticulum stress experimental study genome-wide inhibitor insight mRNA Translation misfolded protein molecular pathology mutant protective pathway protein folding response ribosome profiling therapeutic development therapeutic target transcription factor translation factor

项目摘要

PROJECT SUMMARY Autosomal Dominant Retinitis Pigmentosa (adRP) is a heritable retinal degeneration disorder that results in progressive vision loss and subsequent blindness. Nearly a third of adRP patients possess a mutation in the light-sensing G-protein coupled receptor, Rhodopsin. Many of these mutations cause the rhodopsin to misfold in the Endoplasmic Reticulum (ER) resulting ER dysfunction that leads to retinal degeneration. The age-related nature of the disease is likely because ER stress response pathways that protect against misfolded rhodopsins decline with age. A better understanding of these pathways may therefore contribute to therapeutic strategy development against adRP and other ER stress-mediated maladies. Here, I focus on one particular ER stress response pathway that is initiated by a transmembrane kinase, PERK. Upon sensing misfolded proteins, PERK phospho-inactivates a translation initiation factor, eIF2α, which inhibits protein synthesis and reduces ER burden. Interestingly, these inhibitory conditions stimulate the translation of a transcription factor, ATF4, due to its unusual 5'UTR. Our understanding of ATF4 induction is largely based on studies in yeast and remains incomplete, thus it remains possible that there are as yet unidentified translation regulators that specifically affect ATF4 translation but not canonical mRNA translation. In addition to translation regulation by phospho- eIF2α, the PERK pathway engages a second translational inhibition mechanism via 4E-BP, which is a direct transcriptional target of ATF4. With two translational inhibition mechanisms being activated by PERK signaling, how are stress responsive transcripts (that are required to ameliorate ER stress) translated? This proposal aims to address this major unanswered question regarding the PERK/ATF4 pathway by employing a Drosophila model of adRP to determine the pathological consequence of ATF4 signaling wherein mutant Rhodopsin-1 (Rh1G69D) imposes ER stress and leads to retinal degeneration. Preliminary studies were conducted by screening RNAi lines targeting various translation initiation factors for loss of an ATF4 reporter activity. This lead to the identification of a poorly characterized translational initiation factor as an unexpected regulator of ATF4. Experiments in cultured mouse embryonic fibroblasts (MEFs) shows a phylogenetically conserved role for this new factor in regulating ATF4 translation. Additional preliminary evidence indicates that the newly identified factor regulates translation at the 5' UTR ATF4. Part of the experiments outlined in this proposal is designed to determine how this factor regulates translation of ATF4 utilizing cutting-edge ribosome profiling techniques. The remainder of the proposal details a strategy to understand the role of the second translational inhibitor downstream of PERK, 4E-BP, in retinal degeneration. If realized, this project will significantly further our understanding of translation control during retinal degeneration in addition to providing valuable pharmaceutical targets for ER stress-mediated maladies.

项目摘要常染色体显性视网膜炎色素（ADRP）是一种可遗传的残留变性障碍，导致渐进的视力丧失和随后的失明。近三分之一的ADRP患者在光感应G蛋白偶联受体Rhodopsin。这些突变中有许多导致视紫红质错误折叠在内塑性网状（ER）中，导致残留变性的ER功能障碍。与年龄有关的该疾病的性质可能是因为ER应力反应途径可防止折叠式视紫红蛋白随着年龄的增长而下降。因此，更好地了解这些途径可能有助于理论策略针对ADRP和其他ER应力介导的疾病的发展。在这里，我专注于一种特殊的ER压力由跨膜激酶启动的响应途径，PERK。感觉到错误折叠的蛋白质后，磷酸化灭活的转换起始因子EIF2α，该因子抑制蛋白质合成并降低ER 负担。有趣的是，这些抑制条件刺激了由于它不寻常的5'Utr。我们对ATF4诱导的理解主要基于酵母研究，并且仍然存在因此，不完整影响ATF4翻译，但不影响规范mRNA翻译。除了磷酸化调节 EIF2α，PERK途径通过4E-BP参与第二个翻译抑制机制，这是直接的 ATF4的转录目标。通过两种翻译抑制机制被PERK信号激活，压力响应式转录本如何翻译（可以改善ER应力）？该建议旨在解决有关PERK/ATF4途径的这个重大问题的问题采用ADRP的果蝇模型来确定ATF4信号的病理后果突变的Rhodopsin-1（Rh1g69d）不可能导致视网膜变性。初步研究是通过筛选靶向各种翻译起始因子的RNAi线进行的损失ATF4报告基因活动。这导致识别不当特征的翻译倡议因素是一个意外的因素 ATF4的调节器。培养的小鼠胚胎成纤维细胞（MEF）的实验显示了系统发育该新因素在控制ATF4翻译中的保守作用。其他初步证据表明新确定的因子调节5'UTR ATF4的翻译。其中概述的一部分实验建议旨在确定该因素如何利用尖端核糖体调节ATF4的翻译分析技术。提案的其余部分详细介绍了了解第二个角色的策略 PERK下游的转化抑制剂，4e-bp，残留变性。如果实现的话，这个项目将除了提供视网膜变性期间翻译控制的理解外，还要进一步 ER应力介导的疾病的有价值的药物靶标。