Wildtype or mutant GPR143-EYFP and TYR-ECFP were co-transfected in COS7s. (a) Ratio of emission intensity after:before bleaching. Controls = single transfected COS7s, TYR-ECFP + A2AAR, and ECFP-EYFP fusion. For wtGPR143, photobleaching and relative quantification was performed in intracellular regions. For mtGPR143, portions of plasma membrane were analyzed. (b) FRET efficiency was quantified for co-transfected COS7s. Controls = Single transfected cells, TYR-ECFP + A2AAR, and ECFP-EYFP fusion protein. Data represent means ±SEM of four (co-transfected cells), three (TYR-ECFP + A2AAR) or two control cells. Unpaired Student t-test: * p<0.05, ** p<0.01, ns not significantly different from control. Values refer to limited regions (See Figures S6, S7, S8). A2AAR, adenosine A2A receptor; mt, mutant; TYR, tyrosinase; wt, wildtype.