β-Arrestin-dependent, G Protein-independent ERK1/2 Activation by the β2 Adrenergic Receptor

摘要: Physiological effects of β adrenergic receptor (β2AR) stimulation have been classically shown to result from Gs-dependent adenylyl cyclase activation. Here we demonstrate a novel signaling mechanism wherein β-arrestins mediate β2AR extracellular-signal regulated kinases 1/2 (ERK 1/2) independent G protein Activation ERK1/2 by the expressed in HEK-293 cells was resolved into two components dependent, respectively, on Gs-Gi/protein kinase A (PKA) or β-arrestins. protein-dependent activity rapid, peaking within 2-5 min, quite transient, blocked pertussis toxin (Gi inhibitor) and H-89 (PKA inhibitor), insensitive depletion endogenous siRNA. β-Arrestin-dependent activation slower onset (peak 5-10 min), less robust, but more sustained showed little decrement over 30 min. It sensitive either β-arrestin1 -2 small interfering RNA. In Gs knock-out mouse embryonic fibroblasts, wild-type recruited β-arrestin2-green fluorescent activated toxin-insensitive ERK1/2. Furthermore, mutant (β2ART68F,Y132G,Y219A β2ARTYY), rationally designed based Evolutionary Trace analysis, incapable could recruit β-arrestins, undergo β-arrestin-dependent internalization, activate ERK. Interestingly, overexpression GRK5 -6 increased phosphorylation β-arrestin recruitment, led formation stable receptor-β-arrestin complexes endosomes, agonist-stimulated phospho-ERK1/2. contrast, GRK2, membrane translocation which requires Gβγ release upon activation, ineffective unless it constitutively targeted plasma prenylation signal (CAAX). These findings that can ERK via GRK5/6-β-arrestin-dependent pathway, is coupling.