Developing a general approach to identify the binding locations of adaptor proteins in clathrin cages.

Clathrin-mediated endocytosis (CME) at the plasma membrane is a fundamental cellular process by which molecules are selectively imported into cells and it plays a central role in a wide range of eukaryotic processes. A cellular balance of CME is emerging as an important influence on normal differentiation and development, and oncogenesis. Clathrin is key to the assembly of a protein coat which surrounds the invaginating membrane during CME. Coat formation enables selection of vesicle contents, control of vesicle size and coordination of vesicle formation and detachment from the membrane. The choice of cargo and its cellular destination are governed by collective interactions of its corresponding receptor, clathrin and a diverse group of adaptor proteins. Therefore, clathrin-adaptor protein interactions are key to the cell’s ability to respond to changing conditions. To understand this process, we need to visualise and quantify the nature of clathrin-adaptor protein interactions. Achieving this remains a difficult challenge due to the large size and heterogeneity of clathrin cages and the fact that endocytic adaptor proteins often contain large unstructured domains. We propose to combine SANS and cryo-EM data to obtain orthogonal information about adaptor protein binding positions on clathrin cage structures. Success with this project will demonstrate the value of combining SANS and cryo-EM data and have wide impact throughout the structural biology community.