Decoding the Molecular Choreography of Diverse Vesicle Release Mechanisms

Project Summary The fusion of large dense-core vesicles (LDCVs) with the plasma membrane is essential for the secretion of peptides, hormones, and growth factors in neurons and endocrine cells. This tightly regulated process governs critical physiological functions, including metabolic homeostasis, immune responses, synaptic transmission, appetite regulation, circadian rhythms, and stress management. Disruptions in vesicle fusion are linked to neurological disorders, diabetes, immune dysfunction, eating disorders, depression, addiction, and cancer. Despite its significance, the molecular mechanisms of LDCV release remain poorly understood. The central hypothesis of this proposal is that distinct v-SNARE-containing vesicle populations form unique SNARE configurations, driving different modes of release and fusion pore kinetics. This innovative hypothesis challenges the prevailing view of a single core protein machinery governing vesicle fusion and suggests multiple vesicle types with varying calcium sensitivities. The unresolved debate between full fusion and the kiss-and-run (KR) mechanism has persisted due to technological limitations and the transient nature of these events. Our SLIM technology overcomes these barriers, enabling real-time tracking of vesicle fates and providing insights into the mechanisms behind KR and full fusion. This project will explore vesicle heterogeneity, synchronous and asynchronous exocytosis, and fusion pore kinetics, providing new insights into vesicle release mechanisms. Utilizing advanced technologies and cellular assays, this MIRA research program will address how vesicle heterogeneity dictates different fusion kinetics in LDCVs (Theme 1) and investigate the role of phospholipase in calcium signaling and lipid dynamics at fusion sites, while developing next-generation suspended lipid membrane platforms based on the design principles of organic neuromorphic devices for observing ultrafast fusion kinetics (Theme 2). The technologies we develop in this proposal will benefit the fields of membrane biophysics, cell biology, and single-molecule imaging. Ultimately, uncovering the principles of LDCV release will link intracellular and intercellular signaling with implications for treating metabolic diseases, psychological disorders, and neurodegenerative diseases. The results will impact numerous scientific fields, including cancer, cardiology, metabolism, and neuroscience.