β-Arrestin mediates the export of ciliary GPR161 but not Smoothened together with the BBSome and intraflagellar transport machinery

Abstract

ABSTRACT Specific G-protein-coupled receptors (GPCRs) exist on the ciliary membrane. Hedgehog signaling activation triggers the import of Smoothened into and export of GPR161 from cilia. The BBSome, which comprises eight Bardet–Biedl syndrome (BBS) proteins, mediates GPCR export, together with the intraflagellar transport (IFT) machinery, containing the IFT-A and IFT-B complexes. The absence of any BBSome subunit or IFT27 (also known as BBS19) (an IFT-B subunit) impairs ciliary GPCR export, including that of GPR161. Plasma membrane GPCRs undergo phosphorylation by GPCR kinases (GRKs) and subsequent binding of β-arrestins [β-arrestin1 (ARRB1) and β-arrestin2 (ARRB2)], which is crucial for clathrin-mediated endocytosis. We here confirmed that GPR161 and β-arrestin are accumulated within cilia in the absence of IFT27 or the BBSome, and that ARRB1 and ARRB2 double-knockout impairs GPR161 export. Notably, we found that activation-mimetic β-arrestin mutants can interact with both the BBSome and ciliary GPCRs, and cause constitutive export of GPR161. Moreover, we demonstrated that GRK2 plays a crucial role in GPR161 export. We here propose that phosphorylated GPR161 recruits β-arrestins, converting them into their activated conformation. Activated β-arrestins then interact with the BBSome, which connects them to the IFT machinery to facilitate GPR161 export.