Membrane Protein Trafficking in Photoreceptors

光感受器中的膜蛋白运输

基本信息

批准号：
8204530
负责人：
WOLFGANG BAEHR
金额：
$ 35.76万
依托单位：
UNIVERSITY OF UTAH
依托单位国家：
美国
项目类别：
财政年份：
2008
资助国家：
美国
起止时间：
2008-12-01 至 2013-11-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8204530
关键词：
Abbreviations Address Affect Anabolism Binding Binding Proteins Caenorhabditis elegans Calmodulin-Binding Proteins Cell Maintenance Cell Survival Cell membrane Cellular biology Cilia Co-Immunoprecipitations Complex Cyclic GMP Cytosol Disease Electron Microscopy Endoplasmic Reticulum Etiology Extracellular Domain Filament G protein coupled receptor kinase GRK1 gene Genes Golgi Apparatus Guanylate Cyclase Integral Membrane Protein KRP protein Knock-out Knowledge Light Maintenance Membrane Membrane Protein Traffic Membrane Proteins Membrane Transport Proteins Microtubules Molecular Motors Motor Mus Opsin Partner in relationship Pathway interactions Peripheral Phenotype Phosphotransferases Photoreceptors Phototransduction Physiology Process Proteins Research Retina Retinal Cone Rhodopsin Ribosomes Role Sequence Homology Side Site Sorting - Cell Movement Structure Testing Transducin Transgenes Transgenic Mice Trimethoprim-Sulfamethoxazole Vertebrate Photoreceptors Vesicle abstracting base guanylate cyclase 1 immunocytochemistry kinetosome mutant novel peripherin phosphodiesterase 6 prenyl prevent protein function recombinase research study retinal rods trans-Golgi Network

项目摘要

Abstract. Membrane Protein Trafficking in Photoreceptors. Central to photoreceptor cell biology is an understanding of the sorting and transport of membrane proteins, both integral and peripherally membrane-associated, within the inner segment. Integral membrane proteins are synthesized by ER-associated ribosomes and exported to the Golgi apparatus. Prenylated or acylated peripheral membrane proteins are synthesized in the cytosol and become ER-associated. Vesicles emerge from the trans-Golgi network (TGN) and transport to the base of the cilium where fusion with the cell membrane occurs. Finally, cargo is assembled for intraflagellar transport. Based on our previous results with guanylate cyclase knockouts, we postulate that some peripheral membrane-associated proteins (PMPs) of the phototransduction machinery cotransport with GC-containing vesicles. In GC1/GC2 double knockout rods, PDE6 failed to transport whereas transducin (T) and opsin kinase (GRK1) were unaffected. In GC1-/- cones, the entire phototransduction machinery was absent from the COS suggesting that formation of a large cargo requires GC1, itself an integral membrane protein. Based on deletion of a prenyl binding protein, PrBP/¿ (Pde6d), we discovered that transport of geranylgeranylated PDE6C and farnesylated GRK1 to cone outer segments was blocked while rod PDE6 and rod T transported as expected. Aim 1 of this proposal will address the identification and functional characterization of novel putative prenyl/acyl binding proteins (UNC119 and UNC119B) by gene knockdowns and knockouts. Aim 2 will examine the role of GC1 in post-Golgi transport of cone membrane proteins, emphasizing the importance of the GC1 extracellular domain and GC activity, as well as the role of kinesin-II in intraflagellar transport (IFT). Aim 3 will elucidate the roles of Ca2+-binding proteins, centrin-1 and centrin-2, in intraflagellar transport by rod- and cone- specific knockouts of their genes. These studies will collectively expand our knowledge regarding the protein functions and interactions that participate in photoreceptor maintenance and renewal. Narrative Membrane Protein Trafficking in Photoreceptors Wolfgang Baehr Photoreceptor outer segments are renewed every ten days. Daily renewal of ~10% (about 100 disks) of the outer segment disk stack requires a continuous high rate of biosynthesis, reliable transport, and targeting pathways to replace outer segment proteins. A central question in photoreceptor cell biology concerns post-biosynthetic transport of membrane-associated proteins and the mechanisms of targeting to the outer segments for disk assembly--pathways that are essential for cell maintenance and survival. Otherwise stated, "How does the highly polarized photoreceptor build an outer segment?" The proposed research examines aspects of integral membrane and peripheral membrane protein transport from photoreceptor inner segment (site of biosynthesis) to the outer segment (site of phototransduction). In aim 1, the role of prenyl- or acyl binding proteins will be investigated. In aim 2, we will explore the roles of guanylate cyclase 1 and kinesin-II, a molecular motor involved in intraflagellar transport. Finally in aim 3, the consequences of photoreceptor-specific deletions of centrin- 1 will be studied. Transport of membrane proteins, particularly post-Golgi transport and intraflagellar transport through the cilium, are complex and insufficiently understood in disease etiology.

抽象的。膜蛋白在光感受器中的运输。膜蛋白的分类和转运，包括整体和周围膜相关的膜蛋白内部片段。 Golgi设备。从跨性别网络（TGN）出现了ER的vesict。纤毛与细胞膜融合在一起，最终将货物组装用于flagellar内的运输。基于我们先前使用鸟烯基环化酶敲除的结果，我们假设一些周围膜相关蛋白（PMPS），带有GC插曲的光转移机械共转移 vesict。（grk1）在GC1 - / - 锥中不受影响，整个光转导机器不存在表明形成lage货物需要GC1，这本身是基于整体膜蛋白降低前结合蛋白PRBP/€的缺失（pde6d），我们发现香烷基化的转运 pDE6C和Farnesylated Grk1到锥外段被阻塞，而Rod PDE6和Rod T则作为运输为预期的。通过基因敲低和敲除蛋白质2的蛋白质（UNC119和UNC119B）将检查AIM 2。 GC1在锥膜蛋白的高尔基后运输中的作用，强调了GC1的重要性细胞外域和GC活性，作为flagellar内转运的作用（IFT）阐明Ca2+结合蛋白，中央1和中心运输的作用这些研究的特定敲除将集体扩展参与光感受器维持和更新的功能和相互作用。感光器中的膜蛋白运输 Wolfgang Baehr 每十天更新光感受器的外部段一次。〜10％（约100个磁盘）外部段磁盘堆栈需要连续高生物合成，可靠的运输和靶向途径的速度较高外部段蛋白质。膜相关蛋白和您的生物合成转运靶向磁盘组件外部段的机制 - 途径对于细胞维持和生存至关重要。高度极化的光感受器建立了一个外部段？检查整体膜和周围膜蛋白的各个方面从光感受器内部段（生物合成位点）转运到外部段（光转移的位点）。蛋白质将在AIM 2中进行研究。环化酶1和驱动蛋白-II，一种参与氟法内转运的分子运动。最后，在AIM 3中，中央光感受器规范删除的后果 1将研究膜蛋白。通过纤毛运输和flagellar运输是复杂的，在疾病病因学中不足以理解。