Molecular Basis of Rod and Cone Photoreceptor Outer Segment Structures

项目概要/摘要 本项目研究脊椎动物感光细胞外膜的分子和细胞机制 节（OS）结构，感光细胞生物学中的一个基本未解决的问题。这种基础科学 知识差距严重限制了对人类致盲性疾病的临床理解（和治疗）， 由破坏OS结构的突变引起的动物。各种各样的遗传性视网膜变性 IRD是由破坏OS细胞器结构的突变引起的。因为需要正常的OS结构 对于健康的视力，但还没有被理解，这是至关重要的，以推进知识如何数以百计的 每个杆和锥所需的膜盘被适当地成形和稳定。 由异常操作系统结构触发的IRD的最长期和众所周知的例子是由以下原因引起的： 外周蛋白-2/rds（P/rds）突变。这个建议建立在我们在前一个项目中的演示之上 在此期间，P/rds直接产生膜曲率，以精确地雕刻数百个膜盘 棒和锥形骨水泥中存在边缘。这一进展为一般的 P/rds突变对OS结构的影响，但留下了P/rds如何改变的关键和基本问题， 活性是受调节的，以及P/rds的遗传缺陷如何产生多种疾病表型。我们的中央 假设P/rds功能通过与rom 1和富含谷氨酸异型相互作用来调节 GARPs蛋白质的差异导致视杆细胞和视锥细胞OS盘结构的差异。研究策略 因此，我将确定P/rds如何与rom 1和GARP一起工作，以构建和维护正常的操作系统 结构，并将研究这些蛋白质的意义杆与锥盘架构，以及如何 突变可以破坏细胞器结构，引发特定的视网膜疾病。 利用人类分子遗传学、体外生物化学、细胞内 功能测定和工程脊椎动物模型的原位分析将用于进行 research.具体目标1将阐明异型（rom 1和GARP）相互作用对P/rds的意义 结构、分子功能和OS磁盘体系结构支持。具体目标2将确定差异 P/rds对杆和锥结构和生存能力的贡献。 总之，这些研究有望阐明正常的视杆和视锥OS盘结构是如何产生的， 通过对人类视网膜健康和视力至关重要的光感受器特异性蛋白质来稳定。这项工作 还将为理解P/rds中的致病性突变如何 差异地影响杆和锥以产生IRD的广泛异质性。

Project Summary / Abstract This project investigates the molecular and cellular mechanisms that govern vertebrate photoreceptor outer segment (OS) structure, a fundamental unsolved problem in photoreceptor cell biology. This basic science knowledge gap severely limits clinical understanding of (and treatments for) blinding diseases in humans and animals caused by mutations that disrupt OS structure. A broad variety of inherited retinal degenerations (IRDs) are caused by mutations that disrupt OS organelle structure. Because normal OS structure is required for healthy vision but is not yet understood, it is essential to advance knowledge of how the many hundreds of membranous disks required by each rod and cone are properly shaped and stabilized. The most longstanding and well-known examples of IRDs triggered by abnormal OS structure are caused by mutations in peripherin-2/rds (P/rds). This proposal builds upon our demonstration in the previous project period that P/rds directly generates membrane curvature, to precisely sculpt the hundreds of membranous disk rims present in rod and cone OSs. This advance provides a clear mechanistic explanation for the general effects of P/rds mutations on OS structure, but leaves open the critical and fundamental questions of how P/rds activity is regulated, and how inherited defects in P/rds generate a diversity of disease phenotypes. Our central hypothesis is that regulation of P/rds function by heterotypic interactions with rom1 and glutamic acid-rich proteins (GARPs) contributes to differences between rod and cone OS disk structures. The research strategy will therefore identify how P/rds works together with rom1 and GARPs to build and maintain normal OS structure, and will investigate the significance of these proteins for rod vs. cone disk architectures and how mutations can disrupt organelle structure to trigger particular retinal diseases. An integrated approach using contributions from human molecular genetics, in vitro biochemistry, in cellulo functional assays, and in situ analyses of engineered vertebrate animal models will be utilized to carry out the research. Specific Aim 1 will elucidate the significance of heterotypic (rom1 and GARP) interactions for P/rds structure, molecular function, and support of OS disk architecture. Specific Aim 2 will identify differential contributions of P/rds to rod and cone structures and viability. Overall, these studies are expected to clarify how normal rod and cone OS disk structures are generated and stabilized by photoreceptor-specific proteins that are essential for human retinal health and vision. This work will also provide an improved mechanistic basis for understanding how pathogenic mutations in P/rds can differentially impact rods and cones to produce a broad heterogeneity of IRDs.