C5a-Stimulated Recruitment of β-Arrestin2 to the Nonsignaling 7-Transmembrane Decoy Receptor C5L2

Abstract

C5L2 (or GPR77) is a high-affinity receptor for the complement fragment C5a and its desarginated product, C5a-desArg. Unlike the classical C5a receptor CD88, C5L2 does not couple to intracellular G-protein-signaling pathways but is thought to function as a decoy receptor. The authors show that stimulation of C5L2 with C5a and C5a-desArg induces redistribution of green fluorescent protein—labeled β-arrestin2 to cytoplasmic vesicles. C3a and C3a-desArg were inactive in the β-arrestin translocation assay. Direct interaction of ligand-stimulated C5L2 with β-arrestin was confirmed using a novel β-galactosidase fragment complementation assay. In this assay, C5L2 was labeled with a mutationally altered peptide fragment of β-galactosidase, whereas β-arrestin2 was labeled with a corresponding deletion mutant of the enzyme. Stable transfection of the modified C5L2 and subsequent stimulation with C5a or C5a-desArg restored β-galactosidase activity in a dose-dependent manner. The subnanomolar potency of β-arrestin coupling in the β-galactosidase fragment complementation assay is in agreement with the affinity of the receptor-ligand interaction. C5L2 is the first example of a 7-transmembrane decoy receptor that couples to β-arrestin in a ligand-dependent manner. This observation supports the notion that G-protein-signaling and β-arrestin coupling can be 2 separate activities of 7-transmembrane receptors. Furthermore, the β-arrestin assays described in this article provide methods of screening for selective C5L2 modulators. (Journal of Biomolecular Screening 2009:1067-1075)

Keywords

GPCR complement C5a C5L2 β-galactosidase green fluorescent protein

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