Modulation of Synaptic transmission: Quantitative Analysis of Gβγ specificity to adrenergic α2a receptor and SNARE

Abstract

Modulation of neurotransmitter exocytosis by activated Gi/o coupled G-protein coupled receptors (GPCRs) is a universal regulatory mechanism used both to avoid overstimulation and to influence circuitry. One of the known modulation mechanisms is the Gβγ and soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) interaction. There are 5 Gβ and 12 Gγ subunits, but specific Gβγs activated by a given GPCR and the specificity to effectors in vivo are not known. In this dissertation, a quantitative MRM analysis of neuronal Gβ and Gγ subunits was developed to detect neuronal Gβ and Gγ subunits interaction with receptors and effectors and understand the specificity to α2a adrenergic receptors (α2aARs) and SNARE. The specificity of Gβ and Gγ subunits to α2aARs in both adrenergic (auto α2aARs) and non-adrenergic neurons (hetero α2aARs), and SNARE in presence of epinephrine were identified. Moreover, the microarchitecture of the Gβγ and SNARE interaction was examined by understanding the molecular interaction of Gβγ and SNAP25 and identifying 47 synaptic proteins that may be in complex with Gβγ and SNARE. These studies form the basis for further investigations to determine the effect of G protein specificity on its downstream signaling, identify potential pathophysiological states in which the Gβγ-SNARE interaction may be dysregulated, and yield additional insights into Gi/o-coupled GPCR-mediated regulation of exocytosis.