Gαq Binds Two Effectors Separately in Cells: Evidence for Predetermined Signaling Pathways

G-proteins transduce signals along diverse pathways, but the factors involved in pathway selection are largely unknown. Here, we have studied the ability of Gαq to select between two effectors—mammalian inositide-specific phospholipase Cβ (PLCβ) and phosphoinositide-3-kinase (PI3K)—in human embryonic kidney 293 cells. These studies were carried out by measuring interactions between eCFP- and eYFP-tagged proteins using Forster resonance energy transfer in the basal state and during stimulation. Instead of association of Gαq with effectors through diffusion and exchange, we found separate and stable pools of Gαq-PLCβ and Gαq-PI3K complexes existing throughout the stimulation cycle. These separate complexes existed despite the ability of Gαq to simultaneously bind both effectors as determined by in vitro measurements using purified proteins. Preformed G-protein/effector complexes will limit the number of pathways that a given signal will take, which may simplify predictive models.