Role of PDZ Domain Containing Proteins in Cell–Cell Adhesion and Cell Polarity in the Mouse Lens

Abstract

Abstract: : Purpose:PDZ domain containing proteins are scaffolding molecules known to play a role in cell polarity, cell–cell adhesion and proliferation in Drosophila. Previous studies in our laboratory have shown that expression of the viral oncoprotein E6, which binds to PDZ domain containing proteins, in the mouse lens epithelium leads to increased proliferation and defects in cellular adhesion and differentiation. In this study we determined the association of PDZ domain containing proteins with adherens junctions molecules and if these junctions are destabilized when PDZ protein function is compromised. Methods:Paraffin sections from E18.5 lenses of mice carrying an insertional mutation in dlg1 (dlggt mice) and lenses from P5 or P10 FVB and transgenic mice expressing E6 or E6 Δ146–151, an E6 mutant that cannot bind to PDZ proteins, were used to analyze the localization of Scrib, Dlg, N–cadherin, ZO–1 and ß–actin by immunofluorescence and confocal microscopy. The subcellular localization of N–cadherin, ZO–1 and ß–actin was assessed by western blotting of membrane and cytoplasmic fractions from FVB and transgenic lenses. Results:Scrib and Dlg spatially overlap with adherens junction molecules N–cadherin and ZO–1. Dlg accumulated strongly at the apical and basal tips of the fiber cells where it overlapped with N–cadherin. In the epithelium, ZO–1 was restricted to the apical membrane where it overlapped with both Dlg and Scrib. In the presence of E6, all of these proteins were mislocalized and overlapped inappropriately along the plasma membranes. Localization of ß–actin, which is normally linked to cadherin and ZO–1–containing junctional complexes, was also disrupted. Western blotting for N–cadherin, ZO–1 and ß–actin confirmed the results obtained by immunofluorescence. In lenses from the E6Δ146–151 mice, protein localizations were indistinguishable from that of control mice. Localization of Scrib and ZO–1 was also disrupted in lenses from the dlggt mice. Conclusions:These results show that PDZ domain containing proteins are involved in controlling cell–cell adhesion and cell polarity in the mouse lens and suggest a conserved function cross–species.

Keywords: cell adhesions/cell junctions