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Assembly, quality control, and physiological regulation of membrane trafficking complexes

Membrane protein trafficking in eukaryotes relies on dozens of protein complexes that recruit cargoes, drive vesicle budding, and catalyze vesicle fusion. While the biological roles of these membrane trafficking complexes have been established, our understanding of how they are assembled is still in its early stages. Our group focuses on two assembly pathways: adaptor protein assembly during vesicle budding and SNARE complex assembly during vesicle fusion. We aim to use findings from these two model pathways to establish general principles governing the assembly of membrane trafficking complexes. In addition, we investigate the quality control, physiological regulation, and disease connections of trafficking complex assembly.

1. Chaperone-assisted Adaptor Protein Assembly (CAPA).

Our group discovered the CAPA pathway while identifying endocytic regulators. Adaptor proteins (APs) are heterotetrameric complexes that function on distinct organelles during clathrin-dependent and -independent vesicle budding. There are five APs (AP1-5) in mammalian cells. We found that the assembly of AP2 is assisted by two chaperones: AAGAB and CCDC32. Our current research focuses on unraveling the molecular mechanisms by which AAGAB and CCDC32 facilitate AP2 assembly. Meanwhile, we are searching for chaperones that assist in the assembly of other APs. Ultimately, we aim to obtain a comprehensive molecular understanding of the CAPA pathway.

2. Chaperone-assisted assembly of SNARE complexes and its roles in metabolism and diabetes.

Intracellular vesicle fusion is catalyzed by a class of membrane-bound proteins known as soluble N-ethylmaleimide sensitive factor attachment protein receptors (SNAREs). While SNAREs supply the energy required for membrane fusion, they cannot assemble properly on their own; their assembly depends on dedicated chaperones such as Sec1/Munc18 (SM) proteins. Our team has conducted the first genome-wide CRISPR screens to systematically identify new regulators of regulated exocytosis—a type of vesicle fusion—in mammalian cells. These screens revealed a group of novel regulators involved in SNARE complex assembly. We are currently investigating how these newly identified factors regulate SNARE complex assembly. Another major goal of this project is to understand the roles of SNAREs and their regulators (chaperones) in metabolic regulation and the pathogenesis of metabolic diseases, primarily using insulin-stimulated GLUT4 exocytosis as a model pathway.

Acknowledgements

We are grateful to funding agencies that generously support or supported our work:

National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)

National Institute of General Medical Sciences (NIGMS)

National Institute on Aging (NIA)

American Diabetes Association

Pew Charitable Trust

American Heart Association

University of Colorado Cancer Center

Cancer League of Colorado

Linda Crnic Institute for Down Syndrome

University of Colorado Boulder

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