The ability of proteins to generate membrane curvature is essential for cellular functions, including clathrin-mediated endocytosis (CME). The formation of the coated pit in CME is driven via recruitment of the clathrin triskelia to the membrane surface by adapter proteins that are known to generate steric pressure that drives membrane bending. However, the ability of clathrin to directly contribute to membrane bending itself has remained unclear. In this study, we use quantitative fluorescence microscopy and transmission electron microscopy to examine membrane fission in reconstituted lipid systems. We have used a synthetic affinity tag to assemble clathrin directly to lipid vesicle surfaces in the absence of adaptor proteins to directly interrogate clathrin-membrane interactions. Our results show that clathrin alone bends membranes, promoting the formation of highly curved fission products with diameters below 50 nm diameter. By using pH modulation and divalent cations to tune clathrin’s ability to assemble, we found that clathrin’s ability to drive fission becomes weaker as its assembly strength increases. Corroborating coarse-grain simulations reveal that rapid formation of rigid, large-scale lattices prevents membranes from deforming, whereas highly dynamic assemblies that can readily exchange subunits and switch between hexagons and pentagons are necessary to generate the curvature required for bending and fission. Further experimentation reveals that clathrin can propagate its regulation of membrane curvature through adaptor proteins that are recruited to membranes by PI(4,5)P2 lipids. Specifically, clathrin demonstrates an ability to selectively enhance or inhibit membrane fission depending on the type of adaptor protein with which it interacts. This work elucidates the influence of clathrin’s lattice organization during CME, providing a broader principle for how coats can remodel membranes without direct lipid binding.
Bouzos et al. (Sun,) studied this question.