MOLECULAR BASIS OF NA K ATPASE POLARITY--RP EPITHELIUM

Na-K-ATP酶是介导细胞外基质转运的必需酶， 穿过运输上皮。 在肾脏，消化道和 在外分泌腺中，Na K ATP酶定位于基底外侧的血浆中 膜，而在视网膜色素上皮中，它被发现在 顶端质膜 我们建议研究的分子基础 这种蛋白质和mRNA的差异组织定位 程度. 将大鼠肾脏、肠和RPE Na K ATP酶 用免疫荧光法在半薄冰冻切片上定位， 针对α和β亚单位的单特异性兔抗体。 将使用相同的抗体来比较α的迁移率 和β亚基。 α和β亚基的肽图（包括去糖基化 β亚基）的水平进行比较， 寻找组织特异性差异。 最后， 肾、肠和RPE中的α和β亚基将被 通过北方印迹分析和S1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， 的分子信息负责的阐明 上皮细胞中顶侧和基底侧蛋白质的分选。

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.