Biophysical Society Conference | Estes Park 2023

Membrane Budding and Fusion

Poster Abstracts

48-POS Board 16 UNCLAMPING AND MEMBRANE FUSION ARE CONCURRENT PROCESSES DURING NEUROTRANSMITTER RELEASE

Ioana C Butu 1 ; Zachary McDargh 1 ; Dong An 1 ; Ben O'Shaughnessy 1 ; 1 Columbia University, Chemical Engineering, New York, NY, USA

Synaptic transmission is based on neurotransmitter (NT) release at neuronal synapses, and is accomplished by a specialized machinery that senses presynaptic Ca entry following an action potential, and responds by fusing vesicle and plasma membranes for NT release through a fusion pore. Ca-triggered unclamping and membrane fusion are commonly thought of as distinct, independent processes, accomplished by distinct machinery components: unclamping is the job of the Ca-sensitive clamp Synaptotagmin 1 (Syt) and other components, while the SNARE proteins are the core fusion machinery. However, Ca-uncaging experiments show a cooperativity of release that gradually decreases at low [Ca 2+ ], inconsistent with the prevailing notion that release requires 3-5 Ca 2+ ions to bind Syt, followed by a final fusion step (Lou et al., 2005). The molecular origin of these effects is unknown. Here we developed coarse-grained molecular dynamics simulations that capture the collective behavior of the multicomponent NT release machinery on physiological release timescales. We find fusion is driven by entropic forces among SNARE complexes (SNAREpins) so more SNAREpins catalyze faster fusion. Consequently, unclamping and fusion are concurrent processes: unclamping continuously unfetters increasing numbers of SNARE complexes, so the fusion rate continuously increases. At low [Ca 2+ ], unclamping is rate-limiting, so release rates are heavily dependent on [Ca 2+ ]. At high [Ca 2+ ], unclamping becomes so fast that SNARE-mediated fusion becomes rate-limiting and the release rates plateau, in agreement with experiment.An interesting consequence of the intertwined nature of unclamping and fusion is that the action potential-evoked vesicle release probability (p ves ) increases with more SNAREs or reduced fusion barriers. This explains increased p ves values observed in experiments with constitutively active syntaxin where increased numbers of SNAREpins are present at the fusion site (Acuna et al., 2014).

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