Clathrin mediates membrane fission and budding by constricting membrane pores

Abstract Membrane budding, which underlies fundamental processes like endocytosis, intracellular trafficking, and viral infection, is thought to involve membrane coat-forming proteins, including the most observed clathrin, to form Ω-shape profiles and helix-forming proteins like dynamin to constrict Ω-profiles’ pores and thus mediate fission. Challenging this fundamental concept, we report that polymerized clathrin is required for Ω-profiles’ pore closure and that clathrin around Ω-profiles’ base/pore region mediates pore constriction/closure in neuroendocrine chromaffin cells. Mathematical modeling suggests that clathrin polymerization at Ω-profiles’ base/pore region generates forces from its intrinsically curved shape to constrict/close the pore. This new fission function may exert broader impacts than clathrin’s well-known coat-forming function during clathrin (coat)-dependent endocytosis, because it underlies not only clathrin (coat)-dependent endocytosis, but also diverse endocytic modes, including ultrafast, fast, slow, bulk, and overshoot endocytosis previously considered clathrin (coat)-independent in chromaffin cells. It mediates kiss-and-run fusion (fusion pore closure) previously considered bona fide clathrin-independent, and limits the vesicular content release rate. Furthermore, analogous to results in chromaffin cells, we found that clathrin is essential for fast and slow endocytosis at hippocampal synapses where clathrin was previously considered dispensable, suggesting clathrin in mediating synaptic vesicle endocytosis and fission. These results suggest that clathrin and likely other intrinsically curved coat proteins are a new class of fission proteins underlying vesicle budding and fusion. The half-a-century concept and studies that attribute vesicle-coat contents’ function to Ω-profile formation and classify budding as coat-protein (e.g., clathrin)-dependent or -independent may need to be re-defined and re-examined by considering clathrin’s pivotal role in pore constriction/closure.