Quality control mechanisms against misfolded rhodopsins in Drosophila.

针对果蝇中错误折叠视紫红质的质量控制机制。

基本信息

批准号：
8664498
负责人：
HYUNG D RYOO
金额：
$ 33.8万
依托单位：
NEW YORK UNIVERSITY SCHOOL OF MEDICINE
依托单位国家：
美国
项目类别：
财政年份：
2013
资助国家：
美国
起止时间：
2013-09-01 至 2015-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8664498
关键词：
Affect Age-Years Alleles Amino Acid Substitution Animal Model Attention Binding Biological Biological Assay Blindness Cells Cytoplasm Development Disease Drosophila genus Employee Strikes Endoplasmic Reticulum Endoplasmic Reticulum Degradation Pathway Gene Dosage Gene Expression Genes Genetic Genetic Transcription Goals Hereditary Disease Homologous Gene Human Inborn Genetic Diseases Individual Longevity Mammalian Cell Mediating Messenger RNA Modeling Mutation Outcome Study Pathway interactions Phosphoric Monoester Hydrolases Phosphorylation Process Property Protein Dephosphorylation Proteins Quality Control RNA Binding RNA Interference RNA Recognition Motif RNA Splicing Regulation Retina Retinal Retinal Degeneration Retinitis Pigmentosa Rhodopsin Role Signal Pathway Signal Transduction Stress Testing Therapeutic Ubiquitin Yeasts age related endonuclease endoplasmic reticulum stress fly genome-wide high throughput analysis insight interest multicatalytic endopeptidase complex mutant novel novel therapeutics overexpression phosphatase inhibitor protective effect protein complex protein folding protein misfolding response tool transcription factor

项目摘要

Retinitis Pigmentosa is a group of inherited disorders that show a progressive loss of retinal function. One of the most common causes of Autosomal Dominant Retinitis Pigmentosa (ADRP) are mutations in the rhodopsin gene that disrupt its encoded protein's folding property. Our long-term goal is to understand how cells respond to stress caused by such rhodopsin proteins once they are synthesized in the endoplasmic reticulum (ER). As most cells have robust quality control mechanisms that can help eliminate such misfolded proteins from the ER, a better understanding of these mechanisms may have therapeutic implications. We focus on two specific ER quality control mechanisms that can help suppress retinal degeneration caused by misfolded rhodopsins. First is ER-Associated Degradation (ERAD), which refers to the ubiquitin-mediated degradation of misfolded proteins from the ER. Stimulation of ERAD can suppress retinal degeneration in a Drosophila model for ADRP, but the underlying mechanism remains poorly understood. In addition, ADRP may be suppressed by an intracellular signaling pathway activated by ER-stress, known as the Unfolded Protein Response (UPR). A central branch of the UPR is mediated by the unconventional splicing of xbp1 mRNA in the cytoplasm, leading to the synthesis of an active xbp1 transcription factor. Among the transcription targets of xbp1 include regulators of ERAD. To investigate mechanisms by which ERAD and the UPR suppress retinal degeneration in animal models of ADRP, we plan to use a combination of classical Drosophila genetics, cell biological analysis and high throughput RNAi assays. Specifically, we plan to investigate the precise mechanism by which misfolded rhodopsins are detected by the ERAD machinery and imported into the cytoplasm for degradation. In addition, we plan to study how the xbp1-mediated UPR pathway is regulated. We will test a specific hypothesis where xbp1 mRNA splicing is modulated by a specific phosphatase, and this phosphatase is in turn regulated by a regulatory subunit that binds to xbp1 mRNA. Any new genes or mechanisms identified through this approach will be examined for possible effects on retinal degeneration in a Drosophila model for ADRP, where an endogenous mutation in a rhodopsin encoding gene triggers a dominant form of age-related retinal degeneration. As the fly model shows a striking degree of similarity with the human condition, we believe that a successful outcome of this study may directly influence the development of new strategies against ADRP in humans.

色素性视网膜炎是一组遗传性疾病，显示出视网膜功能的逐渐丧失。常染色体显性视网膜炎色素（ADRP）的最常见原因之一是呈蓝订光蛋白基因中的突变，破坏了其编码蛋白质的折叠特性。我们的长期目标是了解细胞在内质网（ER）中合成的这种视紫红质蛋白引起的压力如何反应。由于大多数细胞具有强大的质量控制机制，可以帮助消除ER中的这种错误折叠的蛋白质，因此对这些机制的更好理解可能具有治疗意义。我们专注于两种特定的ER质量控制机制，可以帮助抑制由错误折叠的视紫红蛋白引起的视网膜变性。首先是与ER相关的降解（ERAD），它指的是泛素介导的蛋白质从ER中脱离错误折叠的降解。刺激ERAD可以抑制ADRP果蝇模型中的视网膜变性，但潜在的机制仍然很少了解。另外，ADRP可以通过ER应力激活的细胞内信号通路抑制，称为展开的蛋白质反应（UPR）。 UPR的中央分支是由XBP1 mRNA在细胞质中的非常规剪接介导的，导致合成活性XBP1转录因子。在XBP1的转录目标中，包括Erad的调节剂。为了研究ADRP动物模型中ERAD和UPR抑制视网膜变性的机制，我们计划使用经典的果蝇遗传学，细胞生物学分析和高吞吐量RNAi分析的组合。具体而言，我们计划研究通过ERAD机械检测到错误折叠的动蛋白的确切机制，并进口到细胞质中以降解。此外，我们计划研究如何调节XBP1介导的UPR途径。我们将检验一个特定的假设，其中XBP1 mRNA剪接由特定的磷酸酶调节，该磷酸酶又受与XBP1 mRNA结合的调节亚基调节。通过这种方法鉴定出的任何新基因或机制，将检查果蝇模型中ADRP视网膜变性的可能影响，其中编码基因的Rhodopsin中的内源性突变触发了与年龄相关的视网膜变性的主要形式。由于苍蝇模型与人类状况相似，因此我们认为这项研究的成功结果可能会直接影响针对人类ADRP的新策略的发展。