CYTOSOLIC MODULATION OF PLASMA MEMBRANE ION TRANSPORT

描述（根据申请人的摘要和具体目标改编）： 项目的重点是质膜钙泵（PMCa）。 PM钙泵是 细胞质钙离子清除的三种主要机制之一 并因此在终止细胞内信号转导中起关键作用。 许多细胞。 钙泵活性的缺陷会导致钙的增加， 随后Na和K发生变化。这些变化将改变关键 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 显示细胞钙稳态的改变。 PMCa提供多个 EP型泵的结构功能研究的优点：与Na PMCa是一个单一的亚单位。 与SR Ca泵相比， 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点。 本申请是 集中于细胞外释放步骤。 IC 50的变化 细胞外Ca抑制可能是由于细胞外Ca ~（2+） 释放步骤、接入通道的改变或移位的变化 步骤或这些步骤的组合。 目标1是确定是否 细胞外Ca释放通过高场进入通道发生。 目的2 是比较细胞外CA和Mg抑制雪貂心脏 当在爪蟾卵母细胞中表达时， 确定改变哪些步骤以说明不同的灵敏度 细胞外Ca.目的3是选择由以下方法产生的抗Ca-out泵： PMCa cDNA的随机诱变（没有细胞的其他改变） 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).