Function of SNAP-23 in H+-ATPase Trafficking in the IMCD

SNAP-23 在 IMCD 中 H-ATP 酶运输中的功能

• 批准号: 6649078
• 负责人: JONATHON H SCHWARTZ
• 金额: $ 5.39万
• 依托单位: BOSTON MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2003
• 资助国家: 美国
• 起止时间: 2003-07-01 至 2004-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6649078
• 关键词: acid base balance; confocal scanning microscopy; exocytosis; green fluorescent proteins; hydrogen transporting ATP synthase; immunocytochemistry; kidney; membrane transport proteins; messenger RNA; postdoctoral investigator; protein binding; protein structure function; protein transport; syntaxin; tissue /cell culture; urine; water channel; western blottings

DESCRIPTION (provided by applicant): The inner medullary collecting duct is a major location of non-volatile acid/base. The IMCD is also responsible for aquaporin- 2 (AQP-2) mediated water reabsorption from the urine. Both the proton pump (H+-ATPase) and AQP-2 are apically targeted in the IMCD cells. In this model of exocytosis, the specific association of a protein(s) on the surface of an H+-ATPase or AQP2 containing vesicle termed v-SNARE (VAMP), and its cognate partner, a membrane protein, termed t-SNARE (Syntaxin). These proteins interact to form a 7S complex, and then, with the interaction of the cytosolic proteins, NSF and SNAP form a 20S fusion complex. Both processes are rate regulated by the balance of exocytic insertion and endocytic retrieval of either the proton pump or the AQP-2 channel. The long term objectives are to elucidate the mechanisms which confer specificity to protein trafficking - which proteins are responsible for the differential regulation of AQP-2 versus H+-ATPase. Further, the role of the ubiquitous protein SNAP-23 has not been elucidated. Utilizing IMCD cell lines in culture, the role of SNAP-23 will be investigated. GFP attached SNAP-23 will be overexpressed in an IMCD cell line. Their response to different pH challenges, compared to wild-type IMCD cells, will be characterized by quantitative mRNA analysis, western blot analysis, immunocytochemical techniques and confocal microscopy. Further, Syntaxin IA and SNAP-23 trafficking and binding characteristics will be examined.

描述（由申请人提供）：内髓集管是不挥发酸/碱的主要位置。IMCD还负责水通道蛋白-2 （AQP-2）介导的尿中水的再吸收。质子泵（H+- atp酶）和AQP-2都是IMCD细胞的靶细胞。在这种胞外作用模型中，含有H+- atp酶或AQP2的囊泡表面的一种蛋白质（称为v-SNARE (VAMP)）与其同源伙伴膜蛋白（称为t-SNARE (Syntaxin)）的特异性结合。这些蛋白相互作用形成7S复合物，然后在胞质蛋白的相互作用下，NSF和SNAP形成20S融合复合物。这两个过程都受到质子泵或AQP-2通道的胞外插入和内吞回收平衡的速率调节。