BIOCHEMISTRY AND MOLECULAR BIOLOGICAL

生物化学和分子生物学

• 批准号: 2160530
• 负责人: MICHELLE M DUFFOURC
• 金额: $ 2.37万
• 依托单位: UNIVERSITY OF SOUTH ALABAMA
• 依托单位国家: 美国
• 财政年份: 1995
• 资助国家: 美国
• 起止时间: 1995-08-21 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/2160530
• 关键词: BIOCHEMISTRY; MOLECULAR; BIOLOGICAL

The general and long term objectives of this research program are to investigate molecules essential to retinal function, and to identify and characterization of a retinal glutamic acid rich protein (garp) which was previously identified in bovine rod outer segments copurifying with rod cGMP phosphodiesterase. More recently, garp has been shown to be covalently linked with the second subunit of the cGMP-gated cation channel of bovine rod photoreceptors, making it a candidate gene for retinal disorders. At least one form of Bardet-Biedl syndrome, a systemic disorder of development that includes retinitis pigmentosa, has been linked to the long arm of chromosome 16. Our finding that the human garp gene is localized to the same region leads us to suggest that defects in garp may be involved in the etiology of this disease. To investigate this possibility, the garp gene will first be characterized in the DNA of normal and Bardet-Biedl patients using heteroduplex and single strand confirmation polymorphism analysis. Any point mutations suggested by these assays will be investigated by direct sequencing of PCR fragments. In conjunction with the gene characterization studies, an immunological characterization of human garp will be undertaken to determine if this protein associates with the channel in an analogous manner to bovine garp. Finally, the expression pattern of garp in neonatal and adult mice will be investigated using ribonuclease protection assay and quantitative Western analysis. Successful completion of these experiments will provide valuable insight into molecular basis of a severe form of retinitis pigmentosa, and provide the information necessary to investigate the role of garp in the function of the cGMP-gated cation channel.

这项研究计划的总体和长期目标是 研究视网膜功能所必需的分子，并识别和 富含谷氨酸的视网膜蛋白(GARP)的特性 以前在牛杆状外节与杆状共同繁殖中被鉴定 CGMP磷酸二酯酶。最近，GARP被证明是 与cGMP门控阳离子的第二亚单位共价连接 牛杆状感光细胞通道，使其成为候选基因 视网膜疾病。至少一种形式的Bardet-Biedl综合征， 包括视网膜色素变性在内的全身性发育障碍 与16号染色体的长臂相连。我们发现人类 GARP基因定位于同一区域，这表明 GARP的缺陷可能与该病的病因有关。至 研究这种可能性，GARP基因将首先被表征 在正常和Bardet-Biedl患者的DNA中使用异源双链和 单链确证多态分析。任何点突变 将通过直接测序的方式进行研究 聚合酶链式反应片段结合基因特征研究， 将对人类GARP进行免疫学表征，以 确定此蛋白质是否与通道以类似的 对待牛的态度。最后，GARP的表达模式在 将使用核糖核酸酶对新生和成年小鼠进行研究 保护实验和定量Western分析。成功 这些实验的完成将为我们提供对 严重形式的视网膜色素变性的分子基础，并提供 研究GARP在GARP功能中的作用所需的信息 CGMP门控阳离子通道。