Biophysical Society Conference | Tahoe 2022

Molecular Biophysics of Membranes

Poster Abstracts

10-POS Board 3 DO MEMBRANES OR SNARE PROTEINS MEDIATE THE EFFECT OF ETHANOL ON VESICLE FUSION? Robert E Coffman 1 ; Alex J Kreutzberger 2 ; Volker Kiessling 3 ; Lukas K Tamm 3 ; Dixon J Woodbury 1 ; 1 Brigham Young University, Neuroscience Center, Provo, UT, USA 2 Harvard University, Cell Biology, Boston, MA, USA 3 University of Virginia Health System, Molec Physiol/Biol Phys, Charlottesville, VA, USA In neurons, neurotransmitter release is highly regulated and sensitive to alcohol. Release of neurotransmitters occurs when synaptic vesicles undergo membrane fusion with the plasma membrane. The SNARE proteins catalyze this reaction by zippering together. The SNARE proteins Syntaxin-1A and SNAP-25 are associated with the plasma membrane and Synaptobrevin-2 is associated with the vesicle membrane. We hypothesize that alcohol may act directly on the fusion machinery or on the fusing membranes, separately from any upstream or downstream process. We used a reconstituted single particle SNARE driven fusion assay (Domanska et al., 2009 JBC. 284:32158-66) to measure vesicle fusion probability with increasing amounts of methanol and ethanol. We also used Circular Dichroism Spectrometry (CD) and site directed fluorescence interference contrast microscopy (FLIC) to measure the effect of these alcohols on neuronal SNARE proteins and their interaction with membranes. We show that fusion probability increases significantly with a physiologically relevant concentration of ethanol (0.4% v/v, 68 mM), but methanol requires a 20-fold higher concentration to see a similar effect. We also show that SNAREs are only affected by alcohol concentrations above 2%. Previously, we reported that ethanol significantly increases fusion rates at 0.4%, in the absence of proteins (Paxman, et al., 2017 Biophys J. 112:121-132). These combined data show that the effect of ethanol on SNARE driven fusion is mediated through its interaction with membranes and not SNARE proteins.

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