2AI activates AhR Signaling in ARPE19 cells (A) 2AI activates luciferase activity (luminescence units on Y axis) of an AhR binding, XRE-luciferase reporter assay in a concentration dependent manner (concentrations displayed on X axis). (B) mRNA levels (from ARPE19 cells) of CYP1a1, an AhR target measured using qPCR treated with varying concentration of 2AI. (C) Comparison of binding of 2AI as compared with the known AhR ligand TCDD from in silico docking. (D) Photo-affinity ligand competition assay using humanized AhR liver cytosol (% photoaffinity label bound the protein) vs. concentrations of 2AI and positive control (BNF). (E) siRNA mediated knockdown of AhR or the untargeted siNT control, followed by relative mRNA expression of AhR targets CYP1a1 and CYP1a2 with or without treatment with 5 μM 2AI (F) mRNA levels of CYP1a1 following treatment with 2AI or co-treatment with AhR antagonist aNF. (n = 3, *p < 0.05).