Release of chemical transmitters by regulated exocytosis underlies many forms of intercellular communication, including hormone release and synaptic transmission. Exocytosis is subject to complex modulation and involves a web of protein-protein interactions and membrane remodeling events. G protein- coupled receptors (GPCRs) play a central role in orchestrating this complex regulation, and in particular are well known to inhibit transmitter release from neurosecretory cells. This profound inhibition contributes to synaptic plasticity and controlling the release of hormones and neuromodulatory peptides. Presynaptic receptors (auto- or hetero-receptors for 5HT, dopamine, acetylcholine, glutamate, etc.) are Gi/o coupled, and they work by the release of G protein beta-gamma subunits. The best-studied mechanism for this inhibition is modulation of the voltage sensitivity of Ca2+ channels. However, Gbeta-gamma can also directly inhibit neurotransmitter release at a point distal to Ca2+ entry by binding to soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) proteins as well as the assembled SNARE complex. The role of this interaction was recently shown by our group to be a physiologically very interesting competitive inhibition of synaptotagmin I (Syt1) binding to the C-terminus of SNAP25. Because this is potentially a very important mechanism for regulation of synaptic activity that may have therapeutic implications, we will pursue the following studies: 1) to determine the structural and molecular basis of Gbeta-gamma binding to SNARE proteins with mutational analysis. 2) We will use these mutants as tools to evaluate the physiological importance of these interactions using transfection, imaging, and amperometric approaches. We previously showed that Gbeta-gamma's inhibition of exocytosis is not due to it's regulation of the classical effectors, but we have recently discovered that Gbeta-gamma binds phospholipase D and inhibits it's activity. Since the product of PLD, phosphatidic acid, is thought to be fusogenic, we will 3) design mutants of Gbeta-gamma and PLD that attenuate their interaction and 4) investigate whether this inhibitory interaction plays a role in the Gbeta-gamma-mediated inhibition of vesicular exocytosis. Our hypothesis of the locus of Gbeta-gamma on SNAREs as a competitive mechanism with synaptotagmin at the exocytotic machinery is elegant, novel, exciting and testable. Thus these studies will have wide implications in neuronal function, as well as secretion in many systems. Because disregulation of presynaptic neurotransmitter function is involved in a large number of diseases, deeper understanding of this interaction may provide future therapeutic targets. ? ? ?

National Institute of Health (NIH)
National Eye Institute (NEI)
Research Project (R01)
Project #
Application #
Study Section
Molecular Neuropharmacology and Signaling Study Section (MNPS)
Program Officer
Mariani, Andrew P
Project Start
Project End
Budget Start
Budget End
Support Year
Fiscal Year
Total Cost
Indirect Cost
Vanderbilt University Medical Center
Schools of Medicine
United States
Zip Code
Yim, Yun Young; Zurawski, Zack; Hamm, Heidi (2018) GPCR regulation of secretion. Pharmacol Ther 192:124-140
Zurawski, Zack; Page, Brian; Chicka, Michael C et al. (2017) G?? directly modulates vesicle fusion by competing with synaptotagmin for binding to neuronal SNARE proteins embedded in membranes. J Biol Chem 292:12165-12177
Van Hook, Matthew J; Babai, Norbert; Zurawski, Zack et al. (2017) A Presynaptic Group III mGluR Recruits G??/SNARE Interactions to Inhibit Synaptic Transmission by Cone Photoreceptors in the Vertebrate Retina. J Neurosci 37:4618-4634
Yim, Yun Young; McDonald, W Hayes; Hyde, Karren et al. (2017) Quantitative Multiple-Reaction Monitoring Proteomic Analysis of G? and G? Subunits in C57Bl6/J Brain Synaptosomes. Biochemistry 56:5405-5416
Zurawski, Zack; Rodriguez, Shelagh; Hyde, Karren et al. (2016) G?? Binds to the Extreme C Terminus of SNAP25 to Mediate the Action of Gi/o-Coupled G Protein-Coupled Receptors. Mol Pharmacol 89:75-83
Hamid, Edaeni; Church, Emily; Wells, Christopher A et al. (2014) Modulation of neurotransmission by GPCRs is dependent upon the microarchitecture of the primed vesicle complex. J Neurosci 34:260-74
Betke, Katherine M; Rose, Kristie L; Friedman, David B et al. (2014) Differential localization of G protein ?? subunits. Biochemistry 53:2329-43
Betke, Katherine M; Wells, Christopher A; Hamm, Heidi E (2012) GPCR mediated regulation of synaptic transmission. Prog Neurobiol 96:304-21
Wells, Christopher A; Betke, Katherine M; Lindsley, Craig W et al. (2012) Label-free detection of G protein-SNARE interactions and screening for small molecule modulators. ACS Chem Neurosci 3:69-78
Wells, Christopher A; Zurawski, Zack; Betke, Katherine M et al. (2012) G?? inhibits exocytosis via interaction with critical residues on soluble N-ethylmaleimide-sensitive factor attachment protein-25. Mol Pharmacol 82:1136-49

Showing the most recent 10 out of 41 publications