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.

项目概要常染色体显性遗传性色素性视网膜炎 (adRP) 是一种遗传性视网膜变性疾病，可导致近三分之一的 adRP 患者存在进行性视力丧失和随后的失明。光感应 G 蛋白偶联受体，视紫红质，其中许多突变会导致视紫红质错误折叠。内质网 (ER) 中的 ER 功能障碍导致视网膜变性。该疾病的性质可能是因为内质网应激反应途径可以防止视紫红质错误折叠因此，更好地了解这些途径可能有助于制定治疗策略。针对 adRP 和其他 ER 应激介导的疾病的发展在这里，我重点关注一种特定的 ER 应激。由跨膜激酶 PERK 感应错误折叠蛋白 PERK 启动的反应途径。磷酸化使翻译起始因子 eIF2α 失活，从而抑制蛋白质合成并减少 ER 暗示，这些抑制条件会刺激转录因子 ATF4 的翻译。我们对 ATF4 诱导的理解主要基于对酵母的研究，并且仍然如此。不完整，因此仍然有可能存在尚未确定的翻译监管机构，专门负责除了通过磷酸化调节翻译外，还影响 ATF4 翻译，但不影响经典 mRNA 翻译。 eIF2α，PERK 通路通过 4E-BP 参与第二种翻译抑制机制，这是一种直接的 ATF4 的转录靶点，通过 PERK 信号激活两种翻译抑制机制，压力反应转录本（改善内质网压力所需的）是如何翻译的？该提案旨在通过以下方式解决有关 PERK/ATF4 途径的这一未解答的主要问题：使用 adRP 的果蝇模型来确定 ATF4 信号传导的病理后果。初步研究表明，突变体 Rhodopsin-1 (Rh1G69D) 会施加 ER 应激并导致视网膜变性。通过筛选针对各种翻译起始因子的 RNAi 系是否丢失 ATF4 报告基因来进行这导致将一个明确表征的翻译起始因子鉴定为意外的。 ATF4 的调节因子在培养的小鼠胚胎成纤维细胞 (MEF) 中的实验显示了系统发育。其他初步证据表明，这一新因子在调节 ATF4 翻译中的保守作用。新发现的因子调节 5' UTR ATF4 的翻译。该提案旨在确定该因子如何利用尖端核糖体调节 ATF4 的翻译提案的其余部分详细介绍了了解第二个角色的策略。 PERK下游的翻译抑制剂，4E-BP，在视网膜变性中的应用如果实现，该项目将除了提供之外，还显着加深了我们对视网膜变性期间翻译控制的理解治疗内质网应激介导的疾病的有价值的药物靶点。