Regulation of Human NHE3 by Ubiquitination

项目总结 腹泻病每年导致近100万名五岁以下儿童死亡。尽管它的发病率是 在较富裕的国家，腹泻的发病率要低得多，腹泻仍然是儿科最常见的两种就诊方式之一 在急诊室，这种情况在住院老人中也很常见。NHE3是一种主要的钠转运体 在小肠和近端结肠的刷状缘膜中。NHE3的异常表达和 功能与急性病原性感染和炎症引起的腹泻疾病有关 我的直觉。本应用程序旨在了解人类NHE3的翻译后修饰。的推动力 这项拟议的研究来自我们最近的发现，人类和灵长类动物的NHE3与人类的NHE3不同 低等哺乳动物，包括啮齿动物和兔子。Nedd4-2是一种与底物相互作用的E3泛素连接酶 通过PY(PPxY)基序的蛋白质。我们发现人类NHE3(HNHE3)与泛素化的Nedd4-2相互作用 HNHE3介导细胞内吞作用，hNHE3对Forskolin/PKA的反应比兔强得多 NHE3.尽管小鼠被广泛地用来了解NHE3的生理作用，但小鼠相对 抗发育性腹泻。我们将考察hNHE3泛素化有助于 人类急性腹泻的严重性增加。首先，我们将比较hNHE3和小鼠的调节 NHE3体内使用转基因动物。我们将研究hNHE3是如何内吞、循环和降解的 Nedd4-2介导的泛素化。泛素化被去泛素化所抵消。我们已经确定了几个 与hNHE3相互作用的假定脱泛酶(DUB)。我们计划调查Ned4-2和Nedd2是如何 DUBS动态调节hNHE3。我们的工作有望揭示新的防治机制。 肠刷与NHE3结合，确定Nedd4-2为治疗腹泻疾病的新靶点 由肠道上皮钠水平衡异常引起。

PROJECT SUMMARY Diarrheal disease causes close to one million deaths in children under five each year. Although its incidence is much lower in the more affluent nations, diarrhea remains one of the two most common visits to pediatric emergency rooms and is also common among the institutionalized elderly. NHE3 is a major sodium transporter in the brush border membrane of the small intestine and proximal colon. Abnormal NHE3 expression and function are associated with diarrheal diseases resulting from acute pathogenic infection and inflammation in the gut. This application aims at understanding post-translational modification of human NHE3. The impetus of the proposed study comes from our recent finding that NHE3s of human and primates differ from NHE3s of lower mammals, including rodents and rabbits. Nedd4-2 is an E3 ubiquitin ligase that interacts with substrate proteins via PY (PPxY) motif. We found that human NHE3 (hNHE3) interacts with Nedd4-2, which ubiquitinates hNHE3 and mediates endocytosis, and the response of hNHE3 is much greater to forskolin/PKA than rabbit NHE3. Although mice have widely been used to understand the physiological role of NHE3, mice are relatively resistant to development diarrhea. We will examine the idea that ubiquitination of hNHE3 contributes to the increased severity of acute diarrhea in man. To begin, we will compare the regulation of hNHE3 and mouse NHE3 in vivo using transgenic animals. We will study how hNHE3 is endocytosed, recycled and degraded by Nedd4-2 mediated ubiquitination. Ubiquitination is counteracted by deubiqutination. We have identified several putative deubiquinating enzymes (DUBs) that interact with hNHE3. We plan to investigate how Nedd4-2 and DUBs dynamically regulated hNHE3. Our work is expected to reveal new mechanism for the control of intestinal brush border NHE3, and identify Nedd4-2 as a novel target for the therapeutic of diarrheal diseases caused by abnormal sodium and water balance in intestinal epithelium.