MOLECULAR BASIS OF NA K ATPASE POLARITY--RP EPITHELIUM

NAKATP酶极性的分子基础--RP上皮

• 批准号: 3426410
• 负责人: Enrique J Rodriguez-Boulan
• 金额: $ 2.28万
• 依托单位: WEILL MEDICAL COLL OF CORNELL UNIV
• 依托单位国家: 美国
• 财政年份: 1987
• 资助国家: 美国
• 起止时间: 1987-08-01 至 1988-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3426410
• 关键词: adenosinetriphosphatase; apical membrane; basolateral membrane; complementary DNA; immunofluorescence technique; messenger RNA; molecular biology; retina; retinal pigment epithelium

Na K ATPase is an essential enzyme for the vectorial transport of Na across transporting epithelia. ln kidney, digestive tract and exocrine glands, Na K ATPase is localized in the basolateral plasma membrane, while in retinal pigmentary epithelium it is found in the apical plasmalemma. We propose to study the molecular basis of this differential tissue localization at the protein and mRNA levels. Rat kidney, intestinal and RPE Na K ATPase will be localized on semithin frozen sections by immunofluorescence using monospecific rabbit antibodies against alpha and beta subunits. The same antibodies will be used to compare the mobility of alpha and beta subunits from these tissues by Western blot analysis. Peptide maps of alpha and beta subunits (including deglycosylated beta subunit) from the three tissues will be compared in order to search for tissue-specific differences. Finally, the mRNAs for alpha and beta subunits in kidney, intestine and RPE will be compared by Northern blot analysis and Sl mapping techniques using cDNAs coding for rat brain alpha and beta subunits. It is expected that these studies will provide important information leading to the, elucidation of the molecular information responsible for the sorting of apical and basolateral proteins in epithelial cells.

Na K ATP酶是载体运输的必需酶 Na 穿过上皮细胞。 在肾脏、消化道和 外分泌腺，Na K ATP酶位于基底外侧血浆 膜，而在视网膜色素上皮中，它存在于 顶端质膜。 我们建议研究其分子基础 蛋白质和 mRNA 的组织定位差异 水平。 大鼠肾、肠和 RPE Na K ATPase 将 通过免疫荧光定位在半薄冰冻切片上 针对 α 和 β 亚基的单特异性兔抗体。 相同的抗体将用于比较 α 的迁移率 通过蛋白质印迹分析从这些组织中分离出β和β亚基。 α 和 β 亚基的肽图（包括去糖基化的 来自三个组织的β亚基）将被比较，以便 寻找组织特异性差异。 最后，mRNA 肾、肠和 RPE 中的 α 和 β 亚基将 通过 Northern blot 分析和 Sl 作图技术进行比较 编码大鼠脑α和β亚基的cDNA。 预计 这些研究将提供重要信息 的，阐明负责的分子信息 上皮细胞中顶端和基底外侧蛋白的分选。