CYTOSOLIC MODULATION OF PLASMA MEMBRANE ION TRANSPORT

描述(根据申请者的抽象和具体目标改编)： 项目的重点是质膜钙泵。粉末冶金钙泵是 清除细胞质钙的三种主要机制之一 因此常常在终止信号转导中起着关键作用 很多细胞。钙泵活性的缺陷会导致钙的增加， 随着Na和K的随后变化，这些变化将改变Key 细胞质功能。高血压，镰状细胞病， 缺血/再灌注损伤、兴奋性毒性、中风和阿尔茨海默病 显示细胞内钙稳态的改变。PMCA提供了几个 EP型泵结构性能研究的优势：不同于Na PUMP，PMCA是一个单亚基。与SR Ca泵不同的是， 蛋白质可以从细胞外介质中获得，这提供了 体细胞选择程序的优势。此应用程序是 重点介绍了细胞外释放的步骤。的IC50的变化 细胞外钙抑制可能是由于细胞外钙的变化引起的 释放步骤、接入通道的改变或移位的改变 步骤或这些步骤的组合。目标1是确定是否 细胞外钙的释放通过高场通道发生。目标2 比较细胞外钙和镁对雪貂心脏的抑制作用 钙泵和大鼠钙泵在非洲爪哇卵母细胞中的表达 确定更改哪些步骤以考虑不同的敏感度 胞外钙离子。目标3是选择由以下方式生成的抗钙出泵 PMCA基因的随机诱变(无其他细胞突变 遗传学)。

DESCRIPTION (Adapted from applicant's abstract and specific aims): The focus of project is the plasma membrane Ca pump (PMCa). The PM Ca pump is one of the three primary mechanisms for the removal of cytoplasmic calcium and thus often plays a pivotal role in terminating signal transduction in many cells. Defects in Ca pump activity would lead to an increase in Ca, with subsequent changes in Na and K. These changes would alter key cytoplasmic functions. Hypertension, sickle cell disease, ischemia/reperfusion injury, excitotoxicity, stroke, and Alzheimer's disease show alterations in cellular Ca homeostasis. The PMCa offers several advantages for structure function studies of EP type pumps: Unlike the Na pump, PMCa is a single subunit. In contrast to the SR Ca pump, parts of the protein are accessible from the extracellular media and this offers advantages for somatic cell selection procedures. This application is focused on the extracellular release steps. Changes in the IC50 for extracellular Ca inhibition could result from changes in the extracellular release steps, alterations of an access channel or changes in translocation steps or a combination of these. Aim 1 is to determine whether extracellular Ca release occurs through a high field access channel. Aim 2 is to compare extracellular CA and Mg inhibition of the ferret cardiac calcium pump with rat calcium pump when expressed in xenopus oocytes and to determine which steps are altered to account for the different sensitivity to extracellular Ca. Aim 3 is to select Ca-out resistant pumps generated by random mutagenesis of the PMCa cDNA (without other alterations of cellular genetics).