Compartmentalized signaling and crosstalk in airway myocytes

PROJECT SUMMARY/ABSTRACT This basic science grant seeks to determine fundamental mechanisms as to how a physiologically relevant cell controls the subcellular location of a signal to optimize the effect of that signal on cell function. We propose to build on recent findings by our diverse investigative team to delineate how 3 different Gs-coupled receptors in airway smooth muscle (ASM) cells - the 2AR, EP2, and EP4 receptors- have their signals compartmentalized within the cell to regulate important ASM cell functions. Aim 1 will employ novel imaging approaches to delineate spatiotemporal features of cAMP/PKA signaling by each receptor and demonstrate how this compartmentalized signaling is shaped by receptor-specific complements of AKAP and PDE isoforms, and by the competing co- activated Gq-coupled receptor. Aim 1 will also employ multiple subcellular-targeted biosensors to characterize the capacity of each receptor to signal from intracellular membrane compartments. Aim 2 will assess how these different receptors generate unique phosphoproteome signatures, and how manipulating the mechanisms shaping localized cAMP/PKA signaling regulates these signatures. Aim 3 will establish how the mechanisms dictating spatiotemporal features of ASM cAMP and the ASM proteome affect Gs-coupled GPCR regulation of ASM contraction, migration, and synthetic functions. The proposed studies will provide a foundation for understanding compartmentalized signaling in the form of both methodological advances and the knowledge gained in how Gs-coupled receptors employ distinct signaling mechanisms to render efficient and specific functional effects. From a translational perspective, our findings will constitute a critical basic science foundation for developing new drugs that target mechanisms of signaling compartments, most readily applied to better control asthma features such as airway hyperresponsiveness, airway remodeling, and possibly airway inflammation.