Negatively curved cellular membranes promote BAIAP2 signaling hub assembly.

Plasma membrane deformations are associated with curvature-dependent protein enrichment that contributes to a wide array of cellular functions. While the spatio-temporal protein dynamics at membrane indentations is well characterized, relatively little is known about protein kinetics at outwardly deforming membrane sites. This is in part due to the lack of high throughput approaches to systematically probe the curvature-dependence of protein-membrane interactions. Here, we developed a nanopatterned array for multiplexed analysis of protein dynamics at negatively curved cellular membranes. Taking advantage of this robust and versatile platform, we explored how membrane shape influences the prototypic negative curvature sensing protein BAIAP2 and its effector proteins. We find assembly of multi-protein signaling hubs and increased actin polymerization at outwardly deformed membrane sections, indicative of curvature-dependent BAIAP2 activation. Collectively, this study presents technical and conceptual advancements towards a quantitative understanding of spatio-temporal protein dynamics at negatively curved membranes.