Diverse pathways in GPCR-mediated activation of Ca mobilization in HEK293 cells.

G protein-coupled receptors transduce extracellular stimuli into intracellular signaling. Ca is a well-known second messenger that can be induced by G protein-coupled receptor activation through the primary canonical pathways involving Gα- and Gβγ-mediated activation of phospholipase C-β (PLCβ). While some G-coupled receptors are shown to trigger Ca mobilization, underlying mechanisms remain elusive. Here, we evaluated whether G-coupled receptors including the β-adrenergic receptor (βAR) and the prostaglandin EP and EP receptors (EPR and EPR) that are endogenously expressed in human embryonic kidney 293 (HEK293) cells utilize common pathways for mediating Ca mobilization. For the βAR, we found an essential role for G in agonist-promoted Ca mobilization while genetic or pharmacological inhibition of G or G had minimal effect. β-agonist-promoted Ca mobilization was effectively blocked by the G-selective inhibitor YM-254890 and was not observed in ΔGα or ΔPLCβ cells. Bioluminescence resonance energy transfer analysis also suggests agonist-dependent association of the βAR with G. For the EPR, which couples to G, agonist treatment induced Ca mobilization in a pertussis toxin-sensitive but YM-254890-insensitive manner. In contrast, EPR, which couples to G and G, exhibited Ca mobilization that was sensitive to both pertussis toxin and YM-254890. Interestingly, both EPR and EPR were largely unable to induce Ca mobilization in ΔGα or ΔPLCβ cells, supporting a strong dependency on G signaling in HEK293 cells. Taken together, we identify differences in the signaling pathways that are used to mediate Ca mobilization in HEK293 cells where the βAR primarily uses G, EPR uses G and G, and EPR uses G, G, and G.