The Biophysical And Biochemical Polarization Of The Epithelial Plasma Membrane

Epithelial cells are structurally and functionally polarized, particularly evident in their compositionally distinct apical and basolateral plasma membrane domains. While extensive research has focused on the distinct protein compositions of these membrane domains, little is known about differences in their lipid composition and resulting membrane biophysical properties. This limitation has persisted due largely to the lack of an effective method for efficiently separating apical from basolateral plasma membranes. Here, we developed a novel technique for the efficient and specific separation of apical and basolateral plasma membranes. By isolating Giant Plasma Membrane Vesicles (GPMVs) from polarized Madin-Darby Canine Kidney (MDCK) cells, we found that the apical and basolateral plasma membranes have distinct biophysical properties. The apical membrane was more tightly packed than the basolateral membrane and had significantly greater domain stability, suggesting robust differences in their lipid compositions. We have extended the GPMV technique to quantify the comprehensive, detailed lipidome of the apical and basolateral plasma membranes, allowing for comparison of differences in lipid head groups, but also in fatty acyl length and unsaturation. Furthermore, we are developing methods to define the compositional and biophysical asymmetry of these membranes. Comparison of the apical and basolateral lipidome reveals novel connections between lipid composition and the biophysical properties of membranes, and more importantly how these properties direct epithelial cell polarization.