Molecular mechanisms of coat assembly and regulation in membrane trafficking pathways

About This Grant

Project Summary The timely delivery of membrane-bound vesicles and tubules bearing transmembrane protein and lipid cargoes to discrete cellular membranes is fundamental to cell biology and human health. Many proteins associated with trafficking pathways are linked to serious and crippling human diseases, especially neurological diseases and disorders. Although many trafficking proteins and some pathways are well characterized, we still do not understand other trafficking pathways that we infer must exist between membranes. This constitutes an enormous gap in our understanding of fundamental cell biology. Our goal is to elucidate the molecular structures and functions of important coat protein complexes that initiate trafficking pathways by forming coats around vesicles or tubules. Coat proteins recognize and package relevant cargoes, and they promote efficient assembly of additional required protein components, like accessory proteins and SNAREs. Clathrin coats have long served as an important paradigm, but increasing evidence demonstrates how other coats use distinct mechanisms. We investigate the retromer and Assembly Polypeptide (AP) family of coat complexes (COPI, AP-4, AP-5) by using a variety of tools to ascertain molecular mechanisms of coat assembly and regulation. Biochemical approaches allow us to identify and test new interactions in coat complexes, including how accessory and regulatory proteins drive function. Integrated structural methods including X-ray crystallography, cryo-electron microscopy (cryo-EM), and cryo-electron tomography (cryo-ET) provide detailed evidence for how coats interact with key partners and allow us to generate specific hypotheses to test function. Biophysical techniques enable us to quantify binding affinities and to probe interfaces identified in structural models. With collaborators, we use molecular data to design experiments in cultured cell lines or in model organisms to explore how protein-protein interactions drive phenotypes at the cellular or organismal levels. Ultimately, we hope to gain a molecular understanding of how coats assemble at distinct membranes to drive different trafficking pathways. We anticipate this work will reveal new mechanisms of coat assembly and regulation and will provide fundamental insights into the protein networks that underlie key cellular events on membranes.