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

• 批准号: 10229185
• 负责人: Deepika Vasudevan
• 金额: $ 6.26万
• 依托单位: NEW YORK UNIVERSITY SCHOOL OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-09-30 至 2021-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10229185
• 关键词: 5' Untranslated Regions; Acute; Address; Affect; Age; Age related macular degeneration; Atherosclerosis; Biochemical; Blindness; Cellular Stress; Chronic; Data; Destinations; Development; Diabetes Mellitus; Disease; Drosophila genus; Embryo; Endoplasmic Reticulum; Etiology; Feedback; Fibroblasts; Functional disorder; G-Protein-Coupled Receptors; Genes; Genetic; Genetic Diseases; Genetic Screening; Genetic Transcription; Genetic Translation; Goals; Heritability; Homeostasis; Impairment; Integral Membrane Protein; Lead; Learning; Light; Mammalian Cell; Mediating; 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 Biosynthesis; Proteins; RNA interference screen; Regulation; Reporter; Research; Retina; 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; base; biological adaptation to stress; design; endoplasmic reticulum stress; experimental study; genome-wide; inhibitor/antagonist; insight; misfolded protein; molecular pathology; mutant; protein folding; response; ribosome profiling; therapeutic development; therapeutic target; transcription factor; translation factor

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色素变性（adRP）是一种遗传性视网膜变性疾病， 进行性视力丧失和随后的失明。近三分之一的adRP患者在基因组中存在突变。 光敏G蛋白偶联受体，视紫红质。许多突变导致视紫红质错误折叠 在内质网（ER）中，导致ER功能障碍，从而导致视网膜变性。年龄相关 这种疾病的性质可能是因为ER应激反应途径，保护对错误折叠的视紫红质 随着年龄的增长而衰退。因此，更好地了解这些途径可能有助于治疗策略 针对adRP和其他ER应激介导的疾病的发展。在这里，我专注于一个特定的ER压力 由跨膜激酶PERK启动的反应途径。在检测错误折叠的蛋白质时，PERK 磷酸灭活翻译起始因子eIF 2 α，其抑制蛋白质合成并减少ER 负担有趣的是，这些抑制条件刺激转录因子ATF 4的翻译，这是由于 不寻常的5 'UTR我们对ATF 4诱导的理解主要基于酵母中的研究， 不完整，因此仍然有可能存在尚未确定的翻译监管机构， 影响ATF 4翻译，但不影响典型mRNA翻译。除了磷酸化的翻译调节外， eIF 2 α，PERK通路通过4 E-BP参与第二种翻译抑制机制，这是一种直接的翻译抑制机制。 ATF 4的转录靶点。由于两种翻译抑制机制被PERK信号转导激活， 如何翻译应激反应转录本（需要改善ER应激）？ 该提案旨在通过以下方式解决关于PERK/ATF 4途径的这一主要未回答的问题： 采用adRP的果蝇模型来确定ATF 4信号传导的病理结果，其中 突变型视紫红质-1（Rh 1G 69 D）施加ER应激并导致视网膜变性。了初步研究 通过筛选靶向各种翻译起始因子的RNA干扰株系， 活动这导致鉴定一个不好的翻译起始因子作为一个意想不到的 ATF 4的调节剂。在培养的小鼠胚胎成纤维细胞（MEFs）中的实验显示， 这一新因子在调节ATF 4翻译中的保守作用。其他初步证据表明， 新鉴定的因子调节5 ′ UTRATF 4的翻译。本文概述的部分实验 一项提案旨在确定该因子如何利用尖端核糖体调节ATF 4的翻译 剖析技术。该提案的其余部分详细介绍了一项战略，以了解第二个 翻译抑制剂下游的PERK，4 E-BP，在视网膜变性。如果实现，该项目将 显著地进一步加深了我们对视网膜变性期间的翻译控制的理解， ER应激介导的疾病的有价值的药物靶点。