Trends in Biochemical Sciences
The ins and outs of G protein-coupled receptor trafficking
Section snippets
Role of phosphorylation in GPCR trafficking
Many GPCRs are targeted for phosphorylation by GRKs, a family of serine/threonine protein kinases that specifically phosphorylate agonist-activated GPCRs 1, 2. GRK-mediated receptor phosphorylation promotes the binding of arrestins that, in turn, promote association of the receptor with components of the endocytic machinery. A classic example of this is the β2AR, which undergoes agonist-dependent phosphorylation by GRK2, binding of arrestin and subsequent endocytosis. Overexpression of
Role of arrestins in GPCR trafficking
A role for arrestins in GPCR trafficking was first suggested by the finding that overexpression of arrestin-2 or -3 promoted internalization of a mutant β2AR impaired in agonist-induced phosphorylation and internalization [14]. Involvement of arrestins in receptor endocytosis was also demonstrated by the inhibitory action of various mutant arrestins on β2AR internalization [14]. Moreover, antisense [15], knockout [16] and RNAi [17] strategies have provided definitive evidence of a direct role
Molecular mechanisms underlying arrestin-promoted GPCR internalization
Initial molecular insight into the mechanism of arrestin-promoted receptor internalization was provided by the finding that non-visual arrestins directly interact with clathrin [13]. The clathrin-binding domain in arrestin-2 and -3 has been localized to a C-terminal clathrin box (Leu-φ-Xaa-φ-[Asp/Glu], where φ represents an aliphatic residue) that is common to a large number of clathrin-binding proteins. Deletion of the clathrin box completely abrogates clathrin binding and reveals an essential
Non-visual arrestin interactions with additional proteins
Non-visual arrestin interaction with several additional proteins also appears to contribute to GPCR trafficking. Non-visual arrestins have been demonstrated to bind ARF nucleotide-binding site opener (ARNO), an ARF guanine-nucleotide-exchange factor and the GDP-bound form of ARF6 [8]. The involvement of ARF6 and ARNO in GPCR regulation was first suggested by the fact that activation of ARF6 by ARNO promotes the release of membrane-bound arrestin-2 and the subsequent desensitization of LH/CG
Additional clathrin-dependent pathways of internalization
Several studies have suggested an arrestin-independent pathway in which GPCRs are internalized through CCPs 50, 51. Studies examining the trafficking of PAR1 in mouse embryonic fibroblasts (MEFs) derived from wild-type, and arrestin-2 and -3 knockout mice have revealed that arrestins are not required for PAR1 internalization, but arrestins do play a role in the desensitization of PAR1 [50]. This finding raises the intriguing possibility that an adaptor other than arrestin fulfils the role of
Post-internalization trafficking
Once plasma membrane proteins are internalized into endocytic vesicles, they are subject to one of two sorting fates (Fig. 2). One fate is the recycling pathway that restores them to the plasma membrane resulting in functional resensitization of receptor-mediated signaling. Another fate is the degradative pathway during which receptors are transported to lysosomes and proteolyzed leading to long-term attenuation of signaling, a process known as down-regulation. In general, GPCRs can be divided
Future directions
Although many recent studies have shed light on the molecular mechanisms mediating GPCR internalization and sorting, many questions remain. Are there additional adaptor molecules that serve to recruit GPCRs to CCPs? If so, what is the molecular composition of these CCPs, and are they functionally distinct from other CCPs? How broad a role does ubiquitination play in sorting GPCRs to the degradative pathway? What are the components of the transport machinery mediating ubiquitin-dependent sorting
Acknowledgements
We thank Raymond Penn and James Keen for their suggestions. This work was supported by grants from the National Institutes of Health (J.L.B. and C.C.) and by postdoctoral fellowships from the Canadian Institutes of Health Research (A.M.) and the American Heart Association (A.M.).
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