| The establishment and maintenance of cell polarity within various organs is crucial for the proper development of both invertebrates and vertebrates. Epithelial cells are polarized in such a way that the apical surface faces a lumen or external environment and the basolateral membrane contacts the substratum and internal compartments. The selective targeting and retention of proteins at either the apical or basolateral surfaces are required for proper epithelial function. In mammalian epithelia, the tight junction is a specialized site of cell contact that acts as a molecular "fence" to separate the apical from basolateral surfaces.; The molecules involved in building epithelial cell architecture have been intensely investigated during recent years and a significant number of studies have shed light on the diverse protein families responsible for orchestrating epithelial polarization. In Drosophila, Crumbs, Stardust, and Discs Lost play a crucial role during epithelial polarization. In this work, the identities of the mammalian orthologues of these proteins are described (CRB3, Pals1, and PATJ, respectively). Biochemical experiments described herein have characterized the protein-protein interaction domains that are responsible for stabilizing the CRB3/Pals1/PATJ complex. Furthermore, the mechanisms by which Pals1 and PATJ localize to tight junctions have been analyzed. Specifically, PATJ serves to target Pals1 to tight junctions. On the other hand, the PATJ/Pals1 association is not sufficient for recruiting PATJ to tight junctions. Instead, this work has demonstrated that a known tight junction protein, ZO-3, could serve to target PATJ to tight junctions. CRB3, which exists in a complex with Pals1 and PATJ, is shown to localize to tight junctions as well as the apical surface suggesting that CRB3, like Drosophila Crumbs, could function as an apical polarity determinant. Consistent with this, the CRB3/Pals1 interaction is shown to be important during the biogenesis of tight junctions and the establishment of apico-basal polarity in mammalian epithelia. |
