ANALYSIS OF ERYTHROCYTE SODIUM-PHOSPHATE COTRANSPORTER

描述(改编自申请人的摘要)：本项目的目标 是为了了解钠-磷酸共转运的分子机制和 钠锂反转运。Na-PO4共转运是一种重要的 通过主动运输对所有哺乳动物细胞至关重要的生理过程 无机磷酸盐，一种中心代谢物。纳力反运输是一种 医学上重要的将锂从细胞中排出的细胞机制。它的 红细胞率与LI治疗效果呈负相关 在双相情感障碍中，并与基础疾病的发展直接相关。 高血压。尽管它们很重要，但人们对这些过程并没有很好的理解 因为负责的分子尚未确定或尚未确定 在模型系统中有充分的特征。最近，在以下方面取得了进展 这两个领域。有间接的动力学和药理学 有证据表明“大脑特有”基因BNP1(或相关的亚型)是 K562红白血病细胞和红细胞Na-PO4共转运体的研究这个 该项目将检验三个假设。假设一：BNP1是红细胞 Na-PO4共转运蛋白。这一假设将在具体目标1中得到检验 证明BNP1是人类红细胞生成的主要亚型 细胞和K562细胞，并在特异靶2中证明BNP1蛋白 存在于这些细胞和成熟的红细胞中。假说二： 红细胞Na-磷酸共转运体也是主要的Na-Na交换器。这 假设将在特定的目标3中通过确定化学计量比来检验 Na-PO4共转运和特异靶4通过表征新的Na转运 BNP1在卵母细胞和HEK293细胞中的异源表达 使用一种新的可诱导启动子系统。假设III：Na-PO4共转运体 是长期以来寻找的Na-Li反向转运的分子基础。这一假设 将在特定的目标5通过确定新的Na-Li反转运进行测试 在表达BNP1的卵母细胞和HEK293细胞中表达。

DESCRIPTION (Adapted from the Applicant's Abstract): The goal of this project is to understand the molecular mechanism of sodium-phosphate cotransport and sodium-lithium countertransport. Na-PO4 cotransport is an important physiological process vital to all mammalian cells through its active transport of inorganic phosphate, a central metabolite. Na-Li countertransport is a medically important cellular mechanism that pumps lithium out of cells. Its rate in erythrocytes inversely correlates with the effectiveness of Li therapy in bipolar disease and directly correlates with the development of essential hypertension. Despite their importance, these processes are not well understood because the responsible molecules have not been identified or have not been adequately characterized in model systems. Recently there has been progress in both of these areas. There is circumstantial kinetic and pharmacological evidence that the "brain specific" gene, BNP1 (or a related isoform), is the Na-PO4 cotransporter in K562 erythroleukemic cells and erythrocytes. The project will test three hypotheses. Hypothesis I: BNP1 is the erythrocyte Na-PO4 cotransporter. This hypothesis will be tested in Specific Aim 1 by demonstrating that BNP1 is the major isoform present in human erythropoietic cells and K562 cells and in Specific Aim 2 by demonstrating that BNP1 protein is present in those cells and mature erythrocytes. Hypothesis II: the erythrocyte Na-phosphate cotransporter is also the major Na-Na exchanger. This hypothesis will be tested in Specific Aim 3 by determining the stoichiometry of Na-PO4 cotransport and in Specific Aim 4 by characterizing the new Na transport caused by the heterologous expression of BNP1 in oocytes and in HEK293 cells using a new inducible promoter system. Hypothesis III: the Na-PO4 cotransporter is the long sought molecular basis for Na-Li countertransport. This hypothesis will be tested in Specific Aim 5 by determining the new Na-Li countertransport in BNP1 expressing oocytes and HEK293 cells.