Tight Junction Proteins and Epithelial Polarity

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DESCRIPTION (provided by applicant): Proper apico-basal epithelial polarity is crucial for normal kidney function and is perturbed in kidney diseases such as acute tubular necrosis and polycystic kidney disease. Recent advances by our group and others have provided new insights into the role of multi-protein complexes in epithelial cell polarization and protein targeting. In the current funding period our laboratory identified scaffolding proteins that localize to the tight junction and are crucial for cell polarization. Our work has focused on proteins associated with the small PDZ domain protein, mLin-7. We demonstrated that one of these mLin-7 binding partners called Protein Associated with Lin Seven 1 (PALS1) localizes to tight junctions and complexes with PALS1 Associated Tight Junction Protein (PATJ) and CrumbsS. PALS1 and PATJ are scaffolding proteins that contain PDZ and L27 domains while CrumbsS is a small apical transmembrane protein. Using siRNA and dominant negative proteins we have demonstrated that PALS1 and PATJ are crucial for epithelial cell polarity. We have also found that PALS1 directly interacts with a common polarity cassette consisting of three proteins Par3/Par6/atypical Protein Kinase C. The goal of our research is to understand how PALS1 and PATJ control the initial steps in epithelial cell polarization. We hypothesize that the scaffolding proteins, PALS1 and PATJ move from an intracellular location to mark a spot in polarizing epithelial cells that directs tight junction localization and the transition from apical to basolateral membrane surfaces. The goals of this proposal are to understand how these proteins come to identify this spot in polarizing epithelial cells and how they proceed to recruit other proteins involved in cell polarization. To achieve these goals we will examine the domains of PALS1 AND PATJ that control their movement within MDCK cells. We will also examine which domains of these proteins are important in localization, trafficking, tight junction formation and cell polarization. In addition, we will examine the signal transduction processes that control these events. These studies will shed new light on the processes necessary for early epithelial polarization and have important implication for renal epithelial function in health and disease.