AbstractClathrin-mediated endocytosis (CME) involves membrane-associated scaffolds of the bin-amphiphysin-rvs (BAR) domain protein family as well as the GTPase dynamin, and is accompanied and perhaps triggered by changes in local lipid composition. How protein recruitment, scaffold assembly and membrane deformation is spatiotemporally controlled and coupled to fission is poorly understood. We show by computational modelling and super-resolution imaging that phosphatidylinositol 3,4-bisphosphate [PI(3,4)P₂] synthesis within the clathrin-coated area of endocytic intermediates triggers selective recruitment of the PX-BAR domain protein SNX9, as a result of complex interactions of endocytic proteins competing for phospholipids. The specific architecture induces positioning of SNX9 at the invagination neck where its self-assembly regulates membrane constriction, thereby providing a template for dynamin fission. These data explain how lipid conversion at endocytic pits couples local membrane constriction to fission. Our work demonstrates how computational modelling and super-resolution imaging can be combined to unravel function and mechanisms of complex cellular processes.