SNAREs In The Trafficking Of IMCD H+-Atpase And AQP2

描述（由申请人提供）：IMCD细胞转运质子，由 水通道蛋白2（AQP 2）介导的H2O穿过其顶端， 膜的在我们培养的IMCD细胞系中，这两个过程都是 受调节的胞吐插入和胞吞回收囊泡控制 它们的细胞膜携带H-ATP酶或AQP 2作为货物，但不 两者SNARE蛋白可能部分介导靶向， 这些囊泡与顶膜的融合。但尽管 假设的靶向融合系统的相似性，H-ATP酶的胞吐作用 和AQP 2是独立调节的。问题是一个极地生物 肾上皮细胞靶向两个不同的载物囊泡的顶端 膜利用类似的对接融合蛋白？为了回答这个问题，我们 提出以下假设，这将是目前的重点 建议：1）最低限度的机械，SNAREpin，所需的目标和 H-ATP酶或AQP 2囊泡亚型与顶膜的融合包括 一组不同的v&tSNARE; 2）货物蛋白（H+-ATP酶和AQP 2），每 se，参与调节，靶向和胞外插入其 载体囊泡和3）这些囊泡的调节胞吐作用不仅是 启动，但通过不同的信号级联进行修饰（磷酸化） 囊泡或靶（顶端）膜中的不同蛋白质。这些 假设将在我们的IMCD细胞系中进行评估，IMCD细胞系是一个优秀的模型系统 用于终末髓集合管功能的研究。这些细胞在 培养物表达许多（如果不是全部）调节H+的酸碱转运蛋白， 分泌和血管紧张素调节H20渗透性调节顶端 针对AQP 2。因此，这些研究为确定 导致细胞外分泌的单独调节的细胞过程 两种不同的转运蛋白插入顶膜。

DESCRIPTION (provided by applicant): IMCD cells transport protons, mediated by an H-ATPase, and H2O mediated by aquaporin-2 (AQP2) across their apical membrane. In our cultured line of IMCD cells, both of these processes are controlled by regulated exocytic insertion and endocytic retrieval of vesicles that carry either an H-ATPase or AQP2 as cargo in their membranes, but not both. It is probable that SNARE proteins, in part, mediate the targeting and fusion of these vesicles to the apical membrane. However, despite the similarity of the postulated targeting-fusion system, exocytosis of H-ATPase and AQP2 are independently regulated. The question then is how does a polar renal epithelial cell target two distinct cargo-laden vesicles to the apical membrane utilizing similar docking-fusion proteins? To answer this question, we propose the following hypotheses which will be the focus of the current proposal: 1) the minimal machinery, the SNAREpin, required for targeting and fusion of H-ATPase or AQP2 vesicle subtypes to the apical membrane consists of a distinct set of v & tSNAREs; 2) the cargo proteins (H+-ATPase and AQP2), per se, participate in the regulation, targeting and exocytic insertion of their carrier vesicles and 3) regulated exocytosis of these vesicles is not only initiated but proceeds by different signal cascades that modify (phosphorylate) dissimilar proteins in either the vesicles or target (apical) membrane. These hypotheses will be evaluated in our IMCD cell line, an excellent model system for the study of terminal medullary collecting duct function. These cells in culture express many if not all of the acid-base transporters that regulate H+ secretion and vasoressin regulated H20 permeability regulated by apically targeted AQP2. Thus these studies provide a unique opportunity for determining the cellular processes that leads to the separate regulation of exocytic insertion of two distinct transporters into the apical membrane.