REGULATION OF THE RENAL BETAINE TRANSPORTER

甜菜碱在保护细胞免受高渗中起重要作用 压力。肾髓质通常为高张状态，肾小球滤过率 高渗性的变化取决于水合状态。在一些 病理状态所有组织都会经历高张状态并适应 不利环境使用的机制类似于 肾髓质。甜菜碱转运体的调控在 甜菜碱在肾上皮细胞中的渗透保护性蓄积。 当细胞暴露在高渗环境中时，转录 编码甜菜碱转运蛋白的基因 刺激导致甜菜碱转运体活性增加 甜菜碱积累。甜菜碱转运体也受到监管 由蛋白激酶A和C的激活物翻译后 为了了解细胞如何感知高渗应激，以及细胞如何 信号被传递到转录机器，即 介导转录刺激的转录因子 甜菜碱转运蛋白基因的高渗反应将被克隆。 使用酵母的遗传选择策略、亲和力筛选或 从纯化的蛋白质中获得的氨基酸序列将被使用 目的：克隆转录因子的全长cDNA。针对该病毒的抗体 转录因子将根据cDNAs序列被提出来。 CDNA和抗体探针将被用来研究 转录因子被高张力激活。如果这项规定 处于Northern杂交确定的转录水平， 核连续分析--转录调控的机制 将通过克隆转录因子基因和 搜索调控序列元件。层级监管 蛋白质丰度的变化，亚细胞定位的转移，以及 磷酸化也将使用抗体进行研究。去学习 蛋白激酶如何抑制甜菜碱转运蛋白的活性 蛋白激酶的激活对血管紧张素转换酶变化的影响 甜菜碱的磷酸化及其亚细胞定位 转运蛋白将使用特定的抗体进行研究。

Betaine plays an important role in protecting cells from hypertonic stress. The renal medulla is normally hypertonic and the degree of hypertonicity changes depending on hydration status. In some pathologic states all tissues experience hypertonicity and adapt to the adverse environment using mechanisms similar to those of the renal medulla. Regulation of the betaine transporter is critical in osmo-protective accumulation of betaine in renal epithelial cells. When cells are exposed to a hypertonic environment, transcription of the gene coding for the betaine transporter is markedly stimulated resulting in increased activity of the betaine transporter and betaine accumulation. The betaine transporter is also regulated post-translationally by activators of protein kinases A and C. In order to understand how cells sense hypertonic stress and how the signal is transduced to the transcriptional machinery, the transcription factor that mediates transcriptional stimulation of the betaine transporter gene in response to hypertonicity will be cloned. A genetic selection strategy using yeast, an affinity screening, or amino acid sequence obtained from the purified protein will be used to clone the cDNA for the transcription factor. Antibodies to the transcription factor will be raised based on the cDNA sequence. The cDNA and antibody probes will be used to study how the transcription factor is activated by hypertonicity. If the regulation is at the level of transcription as determined by Northern blot and nuclear run-on assays, the mechanism of transcriptional regulation will be studied by cloning the gene for the transcription factor and searching for regulatory sequence elements. Regulation at the level of protein abundance, shift in subcellular localization, and phosphorylation will be also studied using the antibodies. To learn how protein kinases inhibit activity of the betaine transporter, the effects of activation of the protein kinases on changes in phosphorylation and subcellular localization of the betaine transporter will be studied using specific antibodies.