MEMBRANES AND PHOSPHOLIPIDS IN RENAL HYPERPLASIA

该项目的长期目标是阐明 细胞外钾（K）减少的机制 浓度被肾上皮细胞转化为促有丝分裂信号 细胞 在缺乏钾的饮食中，大鼠的肾脏生长被诱导， 将K返回到食物中。 我们已经证明了细胞 细胞增殖、细胞器生物发生和磷脂生物合成， 在这个实验模型中，细胞膜可以被打开和关闭， 这种简单的饮食控制 用K-培养基喂养大鼠肾脏， 小时表现出增强的糖酵解，糖酵解酶的激活 甘油醛-3-磷酸脱氢酶（G3 PD），并加速生长。 G3 PD的快速激活是由一种胞浆蛋白介导的， 从暴露于低K促有丝分裂剂的BSC-1细胞的胞质溶胶中纯化 发信号并进行酶裂解。 氨基酸微测序 五个肽片段揭示了修饰剂是一种新的 蛋白 最近利用单特异性抗体的研究表明， 免疫反应性蛋白质和修饰剂功能的增加量， 存在于大鼠肾乳头和红髓质内纹， 出现钾耗竭性肾病。 一种新的协议，以快速纯化 使用免疫亲和柱开发了修饰剂， 促进旨在确定这一系统的结构和功能的研究 蛋白 这一修订申请的具体目的是： 1.采用新的策略从猴肾上皮细胞中获得cDNA克隆 细胞或大鼠肾组织，其编码G3 PD修饰剂，并使用它来 预测蛋白质的氨基酸序列; 2.利用该cDNA克隆研究正常人中基因表达的调控， 和增生的肾组织，以及 3.使用单特异性抗体来阐明 修饰蛋白来控制细胞生长。 实现这些目标可以阐明G3 PD修饰蛋白的作用， K在生理和病理状态下的变化。

THe long-term objective of this project is to elucidate molecular mechanisms by which a reduction in extracellular potassium (K) concentration is transduced into a mitogenic signal by renal epithelial cells. Kidney growth is induced in rats fed a diet deficient in K, and can be reversed by returning K to the chow. We have shown that cell proliferation, organelle biogenesis, and phospholipid biosynthesis for new cellular membranes in this experimental model can be turned on and off by this simple dietary manipulation. The kidneys of rats fed a K-medium for 1 hour exhibit enhanced glycolysis, activation of the glycolytic enzyme glyceraldehyde-3-phosphate dehydrogenase (G3PD), and accelerated growth. The rapid activation of G3PD is mediated by a cytosolic protein was purified from the cytosol of BSC-1 cells exposed to the low-K mitogenic signal, and subjected to enzymatic cleavage. Amino acid microsequencing of five of the peptide fragments revealed that the modifier is a novel protein. Recent studies utilizing a monospecific antibody revealed that an increased amount of immunoreactive protein and modifier function are present in rat renal papilla and inner stripe of red medulla during the onset of K depletion nephropathy. A new protocol to rapidly purify the modifier has been developed, using an immunoaffinity column, that will facilitate studies aimed at defining the structure and function of this protein. The specific aims of this revised application are to: 1. use new strategies to obtain a cDNA clone from monkey kidney epithelial cells or rat kidney tissue which encodes the G3PD modifier, and use it to predict the amino acid sequence of the protein; 2. use the cDNA clone to study the regulation of gene expression in normal and proliferation kidney tissue, and 3. use the monospecific antibody to elucidate the contribution of the modifier protein to control of cell growth. Achieving these aims could elucidate the role of the G3PD modifier protein and K in physiological and pathological states.