GIP receptor suppresses PAC1receptor‐mediated neuronal differentiation via formation of a receptor heterocomplex

Abstract

Pituitary adenylate cyclase-activating peptide (PACAP) receptor (PAC1R) is a class B Gprotein-coupled receptor (GPCR) that is widely expressed in the human body and is involved in neuronal differentiation. As class B GPCRs are known to form heterocomplexes with family members, we hypothesized that PAC1R mediates neuronal differentiation through interaction with a class B GPCR. We used the BRET assay to identify potential interactions between PAC1R and 11 class B GPCRs. Gastric inhibitory polypeptide receptor (GIPR) and secretin receptor were identified as putative binding partners of PAC1R. The effect of heterocomplex formation by PAC1R on receptor activation was evaluated with the cyclic (c)AMP, luciferase reporter, and calcium signaling assays; and the effects on receptor internalization and subcellular localization were examined by confocal microscopy. The results suggested he PAC1R/GIPR heterocomplex suppressed signaling events downstream of PAC1R, including cAMP production, serum response element and calcium signaling, and β-arrestin recruitment. Protein–protein interaction was analyzed in silico, and induction of neuronal differentiation by the PAC1R heterocomplex was assessed in SH-SY5Y neuronal cells by measure the morphological changes and marker genes expression by real-time quantitative PCR and western blot. Over-expression of GIPR suppressed PACAP/PAC1R-mediated neuronal differentiation and the differentiation markers expression in SH-SY5Y cells. GIPR regulates neuronal differentiation through heterocomplex formation with PAC1R.