pH Regulation of Fat Storage in the Nematode Intestine

描述（由申请人提供）： Na+/H+或Cl-/HCO 3-交换的缺陷损害了哺乳动物肠道中的液体和电解质转运，但酸碱转运在营养吸收过程和脂质代谢中的最终作用相对知之甚少。此外，关于肠道pH值如何影响响应营养物质可用性的细胞信号传导事件知之甚少。我们假设pHi作为一个协同信使，并提供了一个代谢的背景下，通过不同的细胞和跨细胞信号传导途径的行动进行解释。在这个应用中，我们提供了初步的证据，连接酸碱运输的脂肪储存和相关的信号事件在线虫C。目的1是研究C. elegans中的SLC 4和SLC 26的Cl-/HCO 3-同源物。通过首先评估它们的分布，然后使用重组蛋白表达在原位和使用遗传编码的pH指示剂和RNAi在体内检查功能，来研究线虫。目的2旨在测试肠道pHi的变化如何影响调节脂肪储存的经典细胞信号传导过程，以及相反地，哪些细胞信号传导过程参与传递细胞酸中毒的表型效应。目的3旨在利用遗传，反向遗传和功能基因组测定来鉴定减弱细胞内pH的调节蛋白，以及对细胞内酸化的全局转录响应。该提案的优势在于，它将使用强大的模型系统和新型试剂来监测生物体中的pH值，从一个独特的角度研究脂肪生物学：酸碱平衡。

DESCRIPTION (provided by applicant): Deficiencies in Na+/H+ or CI-/HCO3- exchange compromise fluid and electrolyte transport in the mammalian intestine, but the ultimate role of acid-base transport in nutrient uptake processes and lipid metabolism is relatively poorly understood. In addition, little is known regarding how intestinal pHi influences cell signaling events that response to nutrient availability. We hypothesis that pHi acts as a synergistic messenger, and provides a metabolic context through which the actions of diverse cellular and trans-cellular signaling pathways are interpreted. In this application, we offer preliminary evidence linking acid-base transport to fat storage and related signaling events in the nematode C. elegans, and propose the following: Aim 1 involves characterizing the SLC4 and SLC26 CI-/HCO3- homologs in C. elegans by first assessing their distribution, then examining function both in situ, using recombinant protein expression and in vivo, using a genetically encoded pH indicator and RNAi. Aim 2 is designed to test how changes in intestinal pHi affect classic cell signaling processes that regulate fat storage, and conversely what cell signaling processes are involved in conveying the phenotypic effects of cellular acidosis. Aim 3 is designed to utilize genetic, reverse-genetic, and functional genomic assays to identify regulatory proteins that attenuate intracellular pH, as well as the global transcriptional responses to intracellular acidification. The strength of this proposal is that it will use a powerful model system and novel reagents for monitoring pH in living organisms to study fat biology from a unique perspective: that of acid-base homeostasis.