Fig. 3

Fig. 3

Benzoquinone structures and hPPAR activities. (A) Structures of idebenone, CoQ₁₀ and derivatives. Note that the isoprenoid side chain contains ten isoprenoid subunits, whereas idebenone contains only alkyl units and ends with a polar hydroxyl group. (B) 2 dpf PPARγ LT embryos showing limited GFP expression in a small subset of epidermal cells and in the posterior spinal cord in the absence of exogenous ligand. Treatment with idebenone, CoQ₂ or CoQ₁₀ induces similar patterns of GFP expression in cells of the epidermis, blood, CNS and posterior spinal cord. Decylubiquinone, which carries a methyl group instead of the hydroxyl group, does not activate the receptor. Benzoquinone, CoQ₀, CoQ₁, CoQ₄, CoQ₆, CoQ₈ and CoQ₉ showed no reporter activation. Pure CoQ₁₀ (Sigma), which has weak water solubility, shows strong receptor activation when mixed together with PEG. Overlay pictures of bright-field and GFP (85% transparent) of 2 dpf embryos are shown. Views are lateral with anterior to the left (mean n=20, replicated three times). (C) Idebenone and CoQ₁₀ are partial dual agonists of PPARα and PPARγ. Embryos of the PPARδ fish line show no increased GFP expression in the presence of CoQ₁₀ or idebenone. Overlaid bright-field and GFP images (85% transparent) of 2 dpf embryos are shown. Views are lateral with anterior to the left (mean n=10, replicated three times). (D) HEK293 cells were co-transfected with the GAL4-LBD fusion proteins of hPPARα, hPPARδ and hPPARγ (as indicated) together with a UAS-luciferase reporter. Treatment of the cells with the full agonists GW7647 (hPPARα), CAY10592 (hPPARδ/β) or rosiglitazone (hPPARγ) resulted in EC₅₀ concentrations of 8 nM, 12.8 nM and 48 nM, respectively. Treatment with idebenone resulted in an EC₅₀ of 2.5 µM for PPARα and 3.8 µM for PPARγ. Notably, maximal activation levels were far lower than elicited by the full agonists. Luciferase output was normalized to β-galactosidase to control for transfection efficiency and expressed as normalized luciferase output. Plasmids used were: pCMX, pcDNA3-GAL4-hPPARα, pcDNA3-GAL4-hPPARδ, pcDNA3-GAL4-hPPARγ, UAS-luc, pGEM, pCMX–β-galactosidase. Data represent mean±s.d., n=3 with at least three repeats.