Investigating The Role of G protein βγ Specificity In Modulation of Synaptic Transmission

Abstract

Synaptic transmission is characterized by exocytotic events which mediate the release of chemical transmitters to facilitate neuronal communication. Inhibitory presynaptic GPCRs act as feedback regulators limiting transmitter release from presynaptic terminals via the actions of their Gβγ subunits. Although Gβγ subunits have been shown to regulate exocytosis through direct interaction with the exocytotic machinery, relatively little is known about which G protein heterotrimers exist in vivo, the specificity of this interaction or its physiological consequences. The hypothesis proposed in the present study is that endogenous Gβγ subunits exhibit specificity when interacting with SNARE proteins to modulate synaptic transmission. To address this, efforts were made to examine the expression of different G protein isoforms throughout the CNS, study the functional specificity of α2A adrenergic receptor mediated Gβγ/SNARE interactions, and develop compounds which would allow its modulation. Targeted proteomics studies demonstrated a wide distribution of most G protein isoforms across brain regions and at synaptic terminals with distinct localization patterns observed for different Gβ and Gγ subunits. Investigation into the functional selectivity of SNARE modulation revealed that α2A adrenergic receptors exhibit specificity when interacting with Gβγ subunits as only a subset of Gβ and Gγ isoforms were coimmunoprecipitated with the receptor following stimulation. Further, lead compounds were developed which had an effect on the Gβγ/SNARE interaction, suggesting it may be possible to target this association directly through the use of selective protein-protein modulators. Taken together, the data presented in this study contribute to a better understanding of G protein signaling within the CNS as well as the role of specificity in α2A adrenergic receptor modulation of SNARE function.