β-Arrestin-mediated activation of MAPK by inverse agonists reveals distinct active conformations for G protein-coupled receptors

Abstract

It is becoming increasingly clear that signaling via G proteincoupled receptors is a diverse phenomenon involving receptor interaction with a variety of signaling partners. Despite this diversity, receptor ligands are commonly classified only according to their ability to modify G protein-dependent signaling. Here we show that ␤2AR ligands like ICI118551 and propranolol, which are inverse agonists for Gs-stimulated adenylyl cyclase, induce partial agonist responses for the mitogen-activated protein kinases extracellular signal-regulated kinase (ERK) 1͞2 thus behaving as dual efficacy ligands. ERK1͞2 activation by dual efficacy ligands was not affected by ADP-ribosylation of G␣i and could be observed in S49-cyc ؊ cells lacking G␣s indicating that, unlike the conventional agonist isoproterenol, these drugs induce ERK1͞2 activation in a Gs͞i-independent manner. In contrast, this activation was inhibited by a dominant negative mutant of ␤-arrestin and was abolished in mouse embryonic fibroblasts lacking ␤-arrestin 1 and 2. The role of ␤-arrestin was further confirmed by showing that transfection of ␤-arrestin 2 in these knockout cells restored ICI118551 promoted ERK1͞2 activation. ICI118551 and propranolol also promoted ␤-arrestin recruitment to the receptor. Taken together, these observations suggest that ␤-arrestin recruitment is not an exclusive property of agonists, and that ligands classically classified as inverse agonists rely exclusively on ␤-arrestin for their positive signaling activity. This phenomenon is not unique to ␤2-adrenergic ligands because SR121463B, an inverse agonist on the V2 vasopressin receptor-stimulated adenylyl cyclase, recruited ␤-arrestin and stimulated ERK1͞2. These results point to a multistate model of receptor activation in which ligand-specific conformations are capable of differentially activating distinct signaling partners.

Nuclear β-adrenergic receptors modulate gene expression in adult rat heart

“…One possibility is that the conformational state of the receptor is altered such that ISO now acts as an inverse agonist for the transcriptional pathway. Recent evidence demonstrates that βAR inverse agonists for AC may act as agonists for activation of MAPKs via a β-arrestinmediated pathway [35]. In our case, similar pleiotropic effects of ISO may result from PKBmediated alterations in the receptor or its signalling partners, leading to the inhibition of de novo RNA synthesis, when PKB is inhibited and activation when it remains activatible.…”

Deconvolution of complex G protein–coupled receptor signaling in live cells using dynamic mass redistribution measurements

“…Activation of downstream signaling events of GPCRs is traditionally recorded with assays based on quantification of distinct intracellular second messengers such as Ca 2+ , IP1 and cyclic AMP 5,9-11 and/or translocation of β-arrestin proteins 5,[12][13][14][15][16] . Since GPCRs from different coupling classes typically produce one or more specific second messenger, and may additionally engage non-G protein effectors [17][18][19][20][21] , several assays are needed to obtain quantitative information about each signaling event. Given the spectrum of cellular activities a single receptor may possess and the dependence of efficacy on the signaling effectors ("pluridimensionality of efficacy" 18 ), the need to analyze integrated cellular responses rather than individual components of signaling pathways is becoming increasingly apparent.…”

Interactions between neurotensin receptors and G proteins

“…Recent data indicate that ligands endowed with inverse agonist activity towards the constitutive activity of "classical" G protein-dependent pathways may induce receptor--arrestin interaction and MAPK activation, like agonists [3]. However, by contrast with agonists, activation triggered…”

“…by inverse agonists appears to occur through G protein-independent mechanisms [3]. In this respect, it would be interesting to evaluate whether such differential mechanisms apply to SR48692, NT and levocabastine, which all activate MAPK in CHO cells [21] but behave as agonist, neutral antagonist and inverse agonist, respectively, for constitutive InsP production in COS cells [62].…”