Quality control mechanisms against misfolded rhodopsins in Drosophila.

项目概要/摘要 色素性视网膜炎是一组遗传性疾病，表现为视网膜功能进行性丧失。其中之一 常染色体显性遗传性视网膜色素变性 (ADRP) 的最常见原因是视紫红质突变 破坏其编码蛋白质折叠特性的基因。我们的长期目标是了解细胞如何反应 一旦这些视紫红质蛋白在内质网（ER）中合成，就会产生应激。作为 大多数细胞具有强大的质量控制机制，可以帮助消除此类错误折叠的蛋白质 ER，更好地理解这些机制可能具有治疗意义。我们重点关注两个具体的 ER 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），它是指泛素介导的错误折叠蛋白的降解。 来自 ER 的蛋白质。刺激 ERAD 可以抑制 ADRP 果蝇模型中的视网膜变性， 但其根本机制仍知之甚少。此外，ADRP 可能被抑制 内质网应激激活的细胞内信号通路，称为未折叠蛋白反应 (UPR)。一个 UPR 的中央分支是由细胞质中 xbp1 mRNA 的非常规剪接介导的，导致 活性 xbp1 转录因子的合成。 xbp1 的转录靶标包括 ERAD 的监管者。研究 ERAD 和 UPR 抑制视网膜变性的机制 ADRP动物模型，我们计划结合经典的果蝇遗传学、细胞生物学分析 和高通量 RNAi 测定。具体来说，我们计划研究精确的机制 错误折叠的视紫红质被 ERAD 机器检测到并导入细胞质中进行降解。在 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 其中 xbp1 mRNA 剪接由特定的磷酸酶调节，而该磷酸酶又受到调节 通过与 xbp1 mRNA 结合的调节亚基。通过此发现的任何新基因或机制 将在 ADRP 果蝇模型中检查该方法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引发与年龄相关的视网膜的显性形式 退化。由于果蝇模型显示出与人类状况惊人的相似程度，我们相信 这项研究的成功结果可能会直接影响针对 ADRP 的新策略的制定 人类。

Project Summary/Abstract 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.