FPR2 signaling without β-arrestin recruitment alters the functional repertoire of neutrophils

Abstract

G-protein coupled receptor (GPCR) biased agonism or functional selectivity has become an essential concept in GPCR research over the last years. Receptor-specific biased agonists selectively trigger one signaling pathway over another and induce a restricted/directed functional response. In this study, we aimed to characterize the concept of biased agonism for FPR2, a member of the formyl peptide receptor (FPR) subfamily of GPCRs. We show that the earlier described FPR2-activating pepducin F2Pal₁₀ is a biased FPR2 agonist. The effects of F2Pal₁₀ on neutrophil function differed in several aspects compared to those mediated by WKYMVM, a conventional FPR2-specific peptide agonist. Upon interaction with FPR2 expressed by neutrophils both F2Pal₁₀ and WKYMVM activated the PLC-PIP₂-Ca²⁺ signaling pathway and the superoxide-generating NADPH-oxidase, but only WKYMVM activated the receptor to recruit β-arrestin. The functional consequences linked to a lack of β-arrestin recruitment were further explored, and we demonstrate that FPR2 desensitization occurred independent of β-arrestin. Despite this, reactivation of desensitized receptors achieved through a disruption of the cytoskeleton and through a novel FPR2 cross-talk mechanism with P2Y₂R (the ATP receptor) and PAFR (the receptor for PAF) differed between F2Pal₁₀-desensitized and WKYMVM-desensitized neutrophils. Further, the inability to recruit β-arrestin was found to be associated with a reduced rate of receptor internalization and impaired chemotaxis in neutrophils. In summary, we provide experimental evidence of biased agonism for FPR2 and our data disclose critical roles of β-arrestin in neutrophil chemotaxis and reactivation of desensitized receptors.

Keywords: Biased agonism; Chemotaxis; Desensitization; Formyl peptide receptor; Pepducin; Reactivation; β-Arrestin.