A Novel Allosteric Activator of Free Fatty Acid 2 Receptor Displays Unique G_i-functional Bias*

Abstract

The short chain fatty acid receptor FFA2 is able to stimulate signaling via both G_i- and G_q/G₁₁-promoted pathways. These pathways are believed to control distinct physiological end points but FFA2 receptor ligands appropriate to test this hypothesis have been lacking. Herein, we characterize AZ1729, a novel FFA2 regulator that acts as a direct allosteric agonist and as a positive allosteric modulator, increasing the activity of the endogenously produced short chain fatty acid propionate in G_i-mediated pathways, but not at those transduced by G_q/G₁₁. Using AZ1729 in combination with direct inhibitors of G_i and G_q/G₁₁ family G proteins demonstrated that although both arms contribute to propionate-mediated regulation of phospho-ERK1/2 MAP kinase signaling in FFA2-expressing 293 cells, the G_q/G₁₁-mediated pathway is predominant. We extend these studies by employing AZ1729 to dissect physiological FFA2 signaling pathways. The capacity of AZ1729 to act at FFA2 receptors to inhibit β-adrenoreceptor agonist-promoted lipolysis in primary mouse adipocytes and to promote chemotaxis of isolated human neutrophils confirmed these as FFA2 processes mediated by G_i signaling, whereas, in concert with blockade by the G_q/G₁₁ inhibitor FR900359, the inability of AZ1729 to mimic or regulate propionate-mediated release of GLP-1 from mouse colonic preparations defined this physiological response as an end point transduced via activation of G_q/G₁₁.