Apical-out polarity in epithelial spheroids requires α6β4 integrins, cell proliferation and anchorage independence.

Epithelial cells primarily segregate transmembrane proteins to apical or basal surfaces, establishing apical-basal polarity. In 3D tissues, apical proteins face inwards. Recent work by our group has shown that increased RhoA activation causes epithelial spheroids to invert apical-basal polarity via a collective rearrangement of cells, a process we and others have termed eversion. In this work, we determined that α6β4-integrin-laminin interactions are required for spheroids to evert to apical-out polarity. Additionally, we show that increased cell proliferation and anchorage independence are required to sustain apical-out polarity. We also observed that apical-out spheroids can 'revert' to apical-in polarity through apoptotic cavitation of cells located in the center of the spheroids. This study provides new mechanistic insights into the biochemical and biophysical mechanisms that drive eversion and maintain apical-out polarity, and supports the concept that apical-basal polarity orientation might drive phenotypic switching of epithelia.