Numb is a membrane active clathrin adaptor

Abstract

Clathrin mediated vesicular transport manages transfer of the bulk of membrane proteins and lipids from the plasma membrane trans Golgi network and endosomes Formation of a clathrin coated carrier involves a complex hierarchy of interactions between clathrin and its adaptors as well as numerous accessory factors that orchestrate cargo sorting membrane budding and fission Numb is an evolutionarily conserved clathrin adaptor that manages the uptake of cognate receptors such as Notch and the cholesterol receptor NPC1L1 from the cell surface How Numb sorts these receptors for clathrin mediated endocytosis remains unclear Here using a novel pulldown template to analyse protein interactions on a membrane surface we identify direct binding between Numb and clathrin Analysis with deletion constructs of Numb maps this interaction to a proline rich region which constitutes a novel clathrin protein interaction module Importantly using compositional mimics of the cell surface we find Numb to display a strong tendency to oligomerize and remodel membranes an attribute that requires the N terminal receptor and lipid binding PTB domain Finally real time fluorescence microscopy based assays indicate that Numb oligomers are highly potent in binding and self assembling clathrin on membranes Together these attributes could form the basis for Numb to function in efficient sorting of cognate receptors and budding of nascent clathrin coated pit newline newline