MOLECULAR BASIS OF PROTEIN TRANSPORT IN PHOTORECEPTORS

描述（申请人描述）：本研究的长期目标 应用程序是了解分子机制， 视紫红质羧基端的天然突变， 视杆细胞感光器中的感光蛋白，引起视网膜色素变性（RP）， 遗传性人类视网膜变性疾病。这些突变被认为 干扰视紫红质向视杆细胞的正确转运 外段我们已经发现，Tctex-1，分子马达的轻链， 细胞质动力蛋白，特异性结合视紫红质的羧基末端。 因此，细胞质动力蛋白可能在细胞外基质的转运中起关键作用。 视紫红质本申请的具体目的是研究 Tctex-1在视紫红质转运中的生理作用。第一个目标是 开发阻断正常功能和/或表达的方法， Tctex-1，无论是在细胞培养和光感受器。第二个目标是 使用所得的试剂和技术来检查抑制 Tctex-1在极化上皮细胞视紫红质转运中的功能/表达 细胞模型系统通过使用细胞培养系统收集的信息 对于我们的第三个目标，即 使用转基因小鼠系统在体内阻断Tctex-1功能/表达。这 实验将明确测试Tctex-1的作用，并通过扩展， 细胞质动力蛋白，在指导视紫红质的适当运输， 光感受器最后，我们的第四个目标将是描述潜在的 蛋白磷酸化在Tctex-1功能调节中的作用 各种体外和细胞培养技术。如果成功，这些 实验将大大增加我们对分子机制的理解 指导和调节视紫红质的正确运输， 感光蛋白这一认识将具有重要的临床意义 相关性，因为提出了视紫红质错误定位和 在某些RP病例中，感光细胞死亡。首先，蛋白质， 视紫红质转运所必需的基因可能是新的候选基因， 遗传性视网膜疾病第二，我们可以获得新的见解， 视紫红质错误定位导致细胞死亡的机制。等 这些见解最终可能被证明对设计保护视力的 治疗RP患者。

DESCRIPTION (Applicant's Description): The long-term goals of this research application are to understand the molecular mechanisms by which naturally-occurring mutations in the carboxy-terminus of rhodopsin, the light-sensing protein in rod photoreceptors, cause retinitis pigmentosa (RP), a hereditary human retinal degenerative disease. These mutations are thought to interfere with the correct transport of rhodopsin to the rod photoreceptor outer segment. We have found that Tctex-1, a light chain of the molecular motor cytoplasmic dynein, binds specifically to the carboxy terminus of rhodopsin. Cytoplasmic dynein may therefore play a critical role in the transport of rhodopsin. The specific aims of this application are designed to study the physiologic role(s) of Tctex-1 in rhodopsin transport. The first aim will be to develop approaches to blocking the normal function and/or expression of Tctex-1, both in cell culture and in photoreceptors. The second aim will be to use the resulting reagents and techniques to examine the effects of suppressing Tctex-1 function/expression on rhodopsin transport in a polarized epithelial cell model system. The information gathered from using a cell culture system for rhodopsin trafficking will be valuable for our third aim, which will be to block Tctex-1 function/expression in vivo using a transgenic mouse system. This experiment will definitively test the role of Tctex-1, and by extension, cytoplasmic dynein, in directing the appropriate transport of rhodopsin in the photoreceptor. Finally, our fourth aim will be to characterize the potential role of protein phosphorylation in the regulation of Tctex-1 function using a variety of in vitro and cell culture techniques. If successful, these experiments will greatly increase our understanding of the molecular mechanisms that direct and regulate the correct transport of rhodopsin and possibly other photoreceptor proteins. This understanding will be of significant clinical relevance because of the proposed link between rhodopsin mistargeting and photoreceptor cell death in some cases of RP. First, proteins that are necessary for rhodopsin transport may represent novel candidate genes for hereditary retinal diseases. Second, we may gain novel insights into the mechanisms by which rhodopsin mislocalization leads to cell death. Such insights may eventually prove to be useful for designing sight-preserving therapies for individuals with RP.