Quality control mechanisms against misfolded rhodopsins in Drosophila.

描述（申请人提供）：色素性视网膜炎是一组遗传性疾病，表现为视网膜功能的进行性丧失。常染色体显性色素性视网膜炎（ADRP）最常见的原因之一是视紫红质基因突变，破坏其编码蛋白的折叠特性。我们的长期目标是了解这种视紫红质蛋白在内质网（ER）中合成后，细胞如何对应激作出反应。由于大多数细胞具有强大的质量控制机制，可以帮助消除内质网中错误折叠的蛋白质，因此更好地了解这些机制可能具有治疗意义。我们关注两种特定的内质网质量控制机制，可以帮助抑制由错误折叠的视紫红质引起的视网膜变性。首先是内质网相关降解（ERAD），指的是泛素介导的内质网错误折叠蛋白的降解。在ADRP的果蝇模型中，刺激ERAD可以抑制视网膜变性，但其潜在机制尚不清楚。此外，ADRP可能被内质网应激激活的细胞内信号通路抑制，称为未折叠蛋白反应（UPR）。UPR的一个中心分支是由细胞质中xbp1 mRNA的非常规剪接介导的，导致一个活性xbp1转录因子的合成。xbp1的转录靶点包括ERAD的调控因子。为了研究ERAD和UPR抑制ADRP动物模型视网膜变性的机制，我们计划将经典果蝇遗传学、细胞生物学分析和高通量RNAi分析相结合。具体来说，我们计划研究ERAD机制检测到错误折叠的视紫红质并将其导入细胞质进行降解的确切机制。此外，我们计划研究xbp1介导的UPR通路是如何被调控的。我们将测试一个特定的假设，即xbp1 mRNA剪接由一个特定的磷酸酶调节，而这个磷酸酶又由一个与xbp1 mRNA结合的调节亚基调节。通过这种方法确定的任何新的基因或机制都将在ADRP的果蝇模型中检查对视网膜变性的可能影响，其中视紫红质编码基因的内源性突变触发了主要形式的与年龄相关的视网膜变性。由于果蝇模型显示出与人类状况惊人的相似性，我们相信这项研究的成功结果可能直接影响人类抗ADRP新策略的发展。

DESCRIPTION (provided by applicant): 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. PUBLIC HEALTH RELEVANCE: Mutations in the rhodopsin gene are responsible for approximately a quarter of autosomal dominant forms of Retinitis Pigmentosa (ADRP), a genetic disease that leads to blindness in individuals 40 ~ 60 years of age. Thus, a better understanding of the mechanisms that help the elimination of misfolded rhodopsins may have a direct impact on developing new therapeutic strategies against this disease.