G protein-coupled receptors (GPCRs) mediate the actions of a wide variety of hormones and neurotransmitters to control functions in all mammalian cells. As such, GPCRs are major targets of therapeutics. Individual GPCRs directly couple to distinct complements of heterotrimeric G protein ? and ?? subunits that drive downstream signaling pathways to shape the cellular responses that determine GPCR efficacy. Both G? and G?? subunits interact directly with effectors to produce cellular responses. Our laboratory has focused largely on G?? subunit signaling and has identified a series of small molecules that bind to G?? to ?bias? signaling pathways downstream of GPCRs. These inhibitors have been applied extensively to dissect G?? functions in biology, and to validate G?? as a viable therapeutic target. Most recently we have identified a small molecule that activates G?? subunit signaling without activating G protein ? subunits. We have applied this tool to study signaling in immune cells to reveal exciting and previously unappreciated roles for G?i in chemokine- chemoattractant receptor signaling. In this proposal we will examine in detail how G?? activating molecules function at a molecular level, and explore the mechanistic role for G?i in chemoattractant receptor signaling in the following specific aims.
Aim 1. Determination of the mechanisms of action of 12155 and M119/gallein on G protein subunit interactions and GPCR coupling.
Aim 2. Determining the mechanism(s) for G?iGTP- dependent regulation of cell migration and adhesion.
Aim 3. Analysis of spatiotemporal regulation of G?i activation in chemoattractant-dependent directional cell migration. We believe that these studies will reveal a new molecular target and cellular function for G?i in regulation of integrin function that will have wide implications for chemoattractant/chemokine receptor and integrin regulation.
G protein coupled receptors (GPCRs) are a major class of transmembrane receptors responsible for recognition of a large class of diverse ligands. Here we propose investigation of selective small molecule inhibitors and activators of G protein ?? subunits identified in our laboratory which could be used to inhibit multiple GPCRs and modify actions of existing GPCR directed pharmaceuticals. Results of these experiments will help to validate this alternate approach to modification of signaling pathways downstream of GPCRs that could ultimately lead to development of novel therapeutics.
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