Induced Nanoscale Membrane Curvature Bypasses Clathrin’s Essential Endocytic Function

During membrane trafficking, flat membrane is rapidly remodeled to produce nanometer-scale vesicles. The mechanisms underlying this shape change are not completely understood, but coat proteins such as clathrin are implicated. Clathrin’s ability to bind to membranes of many different geometries casts uncertainty on its specific role in curvature generation and stabilization. Here, we used nanopatterning to produce substrates of ideal optical properties for live-cell imaging, with U-shaped features that bend a cell’s ventral plasma membrane into shapes characteristic of the energetically-unfavorable intermediate of clathrin- mediated endocytosis (CME). This induced plasma membrane curvature recruits the endocytic machinery, promoting productive endocytosis. Upon clathrin or AP2 disruption, CME sites on flat substrates are diminished. However, induced curvature rescues the localization, turnover and transferrin cargo uptake activities of CME sites after clathrin, but not AP2, disruption. These data establish that clathrin’s essential function during CME is to facilitate the evolution of membrane curvature rather than to scaffold CME protein recruitment.