Fig. 1.
4-CMTB is a selective ago-allosteric modulator at FFA2. Flp-In TREx 293 cells stably expressing inducible hFFA2-eYFP (A, C, and D) or hFFA3-eYFP (B) were induced with 0.5 μg/ml doxycycline for 24 h before harvesting and membrane preparation for subsequent [35S]GTPγS incorporation assays. A, both 4-CMTB and propionate are full agonists in this assay. Inset, representative example of absolute values [disintegrations per minute (DPM)] for basal (−) and propionate (C3; 10 mM) and 4-CMTB (10 μM) from a single hFFA2-eYFP membrane preparation. B, 4-CMTB is not an agonist, antagonist, or allosteric modulator of propionate at the closely related hFFA3-eYFP receptor. Inset, representative experiment for hFFA3-eYFP, as per A inset. C, 4-CMTB and propionate interact allosterically, as increasing fixed concentrations of 4-CMTB result in enhanced potency of propionate. The diagonal arrow indicates the shift in potency. *, p < 0.05 according to one-way ANOVA with Dunnett's post hoc analysis. D, the allosteric relationship is reciprocal. Again, the diagonal arrow indicates the shift in potency. *, p < 0.05 according to one-way ANOVA with Dunnett's post hoc test. Data are mean ± S.E.M (n = 3).