GPR3 Stimulates Ab Production via Interactions with APP and b-Arrestin2

The orphan G protein-coupled receptor (GPCR) GPR3 enhances the processing of Amyloid Precursor Protein (APP) to the neurotoxic beta-amyloid (Ab) peptide via incompletely understood mechanisms. Through overexpression and shRNA knockdown experiments in HEK293 cells, we show that b-arrestin2 (barr2), a GPCR-interacting scaffold protein reported to bind c-secretase, is an essential factor for GPR3-stimulated Ab production. For a panel of GPR3 receptor mutants, the degree of stimulation of Ab production correlates with receptor-b-arrestin binding and receptor trafficking to endocytic vesicles. However, GPR3’s recruitment of barr2 cannot be the sole explanation, because interaction with barr2 is common to most GPCRs, whereas GPR3 is relatively unique among GPCRs in enhancing Ab production. In addition to b-arrestin, APP is present in a complex with GPR3 and stimulation of Ab production by GPR3 mutants correlates with their level of APP binding. Importantly, among a broader selection of GPCRs, only GPR3 and prostaglandin E receptor 2 subtype EP2 (PTGER2; another GPCR that increases Ab production) interact with APP, and PTGER2 does so in an agonist-stimulated manner. These data indicate that a subset of GPCRs, including GPR3 and PTGER2, can associate with APP when internalized via barr2, and thereby promote the cleavage of APP to generate Ab