Physiological and pharmacological implications of beta-arrestin regulation

Abstract

G protein-coupled receptor-targeted drug discovery as well as “compound reassessment” requires the utilization of diverse screens to determine agonist efficacies and potencies beyond the scope of ligand binding and G protein coupling. Such efforts have arisen from extensive studies, both in cellular and animal models, demonstrating that these seven transmembrane domain-spanning, G protein-coupled receptors may engage in more diverse functions than their name suggests and particular focus is drawn to their interactions with beta-arrestins (βarrestins). As regulators, βarrestins are involved in dampening G protein-coupling pathways. βArrestins can also play pro-signaling roles in receptor mediated events and the coupling of receptors to βarrestins may be as important as their potential to couple to G proteins in the physiological setting. In the last decade, the development of βarrestin deficient mouse models has allowed for the assessment of the contribution of individual βarrestins to receptor function in vivo. This review will discuss the current literature that implicates βarrestins in receptor function in respect to physiological and behavioral responses observed in the live animal model.

Introduction

βArrestins (non-visual arrestins) are ubiquitously expressed proteins that were first described for their role in desensitizing G protein-coupled receptors (GPCRs). There are two βarrestins, namely, βarrestin1 and βarrestin2, which are also referred to as arrestin-2 and arrestin-3, respectively. As their names imply, βarrestins were first identified for their ability to “arrest” agonist-stimulated β2 adrenergic receptor (β2AR) signaling (Lohse et al., 1990) in a manner similar to arrestin regulation of rhodopsin. The canonical model of GPCR regulation by βarrestins also involves GPCR kinases (GRKs) which phosphorylate receptors and thereby serve to facilitate receptor-βarrestin interactions (Benovic et al., 1987, Sibley et al., 1987, Lohse et al., 1992, Pitcher et al., 1992). Upon complexing with receptors, βarrestins can serve as inhibitors of signal transduction by preventing further receptor coupling to G protein signaling cascades (for reviews see: Premont et al., 1995, Freedman and Lefkowitz, 1996, Lefkowitz, 1998).

Specific examples of βarrestins serving as negative regulators of GPCR signaling are plentiful in cellular as well as animal model systems (Table 1) (for reviews see: Gainetdinov et al., 2004, Bohn et al., 2004a, Gurevich and Gurevich, 2006, Premont and Gainetdinov, 2007). In addition to mediating receptor desensitization, βarrestins can facilitate recruitment and interactions between GPCRs and signaling partners. In this capacity, βarrestins can serve as positive mediators of receptor signaling to downstream targets. Evidence for GPCRs coupling to βarrestins to transduce receptor signaling has also been widely demonstrated in cellular models (for reviews see: Luttrell et al., 1999, Luttrell, 2002, Lefkowitz and Shenoy, 2005, DeWire et al., 2007). Studies in mouse models also support a pro-signaling role for βarrestins (particularly βarrestin2) and these reports are summarized in Table 2.

Arguably, the most studied GPCR is the β2AR. In vitro studies with this receptor have been instrumental in demonstrating the diverse and pleiotropic roles that βarrestins can play in determining agonist-induced receptor responses. The β2AR has been shown to interact with both βarrestin1 and βarrestin2 upon agonist stimulation (Attramadal et al., 1992) and such interactions result in decreased responsiveness to agonist over time. The removal of βarrestins by early anti-sense studies (Mundell et al., 1999), later siRNA studies (Ahn et al., 2003), as well as studies utilizing mouse embryonic fibroblasts devoid of both βarrestin1 and βarrestin2 (Kohout et al., 2001), confirm that βarrestins play a critical role in promoting this waning effect on G protein-coupling and adenylyl cyclase stimulation following agonist activation of the β2AR. Similar studies have been performed for multiple GPCRs of diverse classes and together, these findings support the canonical model wherein the agonist-activated GPCR becomes phosphorylated by GRKs which subsequently increases the binding affinity of the receptor for βarrestins.

βArrestin interactions with activated GPCRs can be detected by co-immunoprecipitation (Groer et al., 2007), confocal microscopy (Barak et al., 1997), bioluminescence resonance energy transfer (BRET) (Hamdan et al., 2005), and fluorescence resonance energy transfer (FRET) (Drake et al., 2008) assays. Such developments, including enzyme complementation assays (von Degenfeld et al., 2007), have facilitated high throughput screens for assessing drug-induced βarrestin-receptor interactions. Looking forward, the interactions between βarrestins and GPCRs may be realized for ultimately determining relative drug efficacies in vivo (Claing and Laporte, 2005, Violin and Lefkowitz, 2007, DeWire et al., 2007).