A key interfacial residue identified with in-cell structure characterization of a class A GPCR dimer

G protein coupled receptors (GPCRs) have been shown homo-dimeric. Despite extensive studies, no single residue has been found essential for dimerization. Lacking an efficient method to shift the monomer-dimer equilibrium also makes functional relevance of GPCR dimer elusive. Here, using fluorescence lifetime-based imaging for distance measurements, we characterize the dimeric structure of GPR17, a class A GPCR, in cells. The structure reveals transmembrane helices 5 and 6 the dimer interface, and pinpoints F229 a key residue, mutations of which can render GPR17 monomeric or dimeric. Using the resulting mutants, we show that GPR17 dimer is coupled to both Gα i and Gα q signaling and is internalized, whereas GPR17 monomer is coupled to Gα i signaling only and is not internalized. We further show that residues equivalent to F229 of GPR17 in several other class A GPCRs are also important for dimerization. Our findings thus provide fresh insights into GPCR structure and function.