PPARα Ligand-Binding Domain Research Articles

14,15-Dihydroxyeicosatrienoic acid activates peroxisome proliferator-activated receptor-α

Epoxyeicosatrienoic acids (EETs), lipid mediators synthesized from arachidonic acid by cytochrome P-450 epoxygenases, are converted by soluble epoxide hydrolase (SEH) to the corresponding dihydroxyeicosatrienoic acids (DHETs). Originally considered as inactive degradation products of EETs, DHETs have biological activity in some systems. Here we examined the capacity of EETs and DHETs to activate peroxisome proliferator-activated receptor-alpha (PPARalpha). We find that among the EET and DHET regioisomers, 14,15-DHET is the most potent PPARalpha activator in a COS-7 cell expression system. Incubation with 10 microM 14,15-DHET produced a 12-fold increase in PPARalpha-mediated luciferase activity, an increase similar to that produced by the PPARalpha agonist Wy-14643 (20 microM). Although 10 microM 14,15-EET produced a threefold increase in luciferase activity, this was abrogated by the SEH inhibitor dicyclohexylurea. 14-Hexyloxytetradec-5(Z)-enoic acid, a 14,15-EET analog that cannot be converted to a DHET, did not activate PPARalpha. However, PPARalpha was activated by 2-(14,15-epoxyeicosatrienoyl)glycerol, which was hydrolyzed and the released 14,15-EET converted to 14,15-DHET. COS-7 cells incorporated 14,15-[3H]DHET from the medium, and the cells also retained a small amount of the DHET formed during incubation with 14,15-[3H]EET. Binding studies indicated that 14,15-[3H]DHET binds to the ligand binding domain of PPARalpha with a Kd of 1.4 microM. Furthermore, 14,15-DHET increased the expression of carnitine palmitoyltransferase 1A, a PPARalpha-responsive gene, in transfected HepG2 cells. These findings suggest that 14,15-DHET, produced from 14,15-EET by the action of SEH, may function as an endogenous activator of PPARalpha.

Inhibition of peroxisome proliferator-activated receptor α signaling by vitamin D receptor

Peroxisome proliferator-activated receptors (PPARs) are nuclear fatty acid receptors that have been implicated to play an important role in lipid and glucose homeostasis. PPARα potentiates fatty acid catabolism in the liver and is activated by the lipid-lowering fibrates, whereas PPARγ is essential for adipocyte differentiation. Here we report that nuclear vitamin D 3 receptor (VDR) represses the transcriptional activity of PPARα but not PPARγ in a 1,25(OH) 2D 3-dependent manner. The analysis using chimeric receptors revealed that ligand binding domain of PPARα and VDR was involved in the molecular basis of this functional interaction and that the DNA binding domain of VDR was not required for the suppression, suggesting a novel mechanism that might involve protein–protein interactions rather than a direct DNA binding. Furthermore, the treatment of rat hepatoma H4IIE cells with 1,25(OH) 2D 3 diminishes the induction of AOX mRNA by PPARα ligands, Wy14,643. VDR signaling might be considered as a factor regulating lipid metabolism via PPARα pathway. We report here the novel action of VDR in controlling gene expression through PPARα signaling.

Corepressor Action

The activity of nuclear receptors such as the peroxisome proliferator-activated receptor-α (PPAR-α), which regulates the expression of genes that modulate fatty acid metabolism, can be regulated through interaction with coactivator and corepressor molecules. Understanding the nature of these interactions may help the development of drugs that effectively control lipid levels. By examining the crystal structure of a ternary complex comprised of the PPAR-α ligand-binding domain, an antagonist molecule, and the receptor-binding motif of the corepressor SMRT, Xu et al. report that a groove in the nuclear receptor contains overlapping binding sites for both coactivators and corepressors. However, in the presence of an antagonist molecule, a specific helix in the receptor is blocked from assuming an active conformation. This block results in a large domain that can specifically accommodate the receptor binding motif of a coprepressor. This mechanism may be utilized by nuclear receptors in general because the corepressor binding sequence is highly conserved.H. E. Xu, T. B. Stnaley, V. G. Montana, M. H. Lambert, B. G. Shearer, J. E. Cobb, D. D. McKee, C. M. Galardi, K. D. Plunket, R. T. Nolte, D. J. Parks, J. T. Moore, S. A. Kliewer, T. M.Willson, J. B. Stimmel, Structural basis for antagonist-mediated recruitment of nuclear co-repressors by PPARα. Nature 415, 813-817 (2002). [Online Journal]

Corepressor Action

The activity of nuclear receptors such as the peroxisome proliferator-activated receptor-α (PPAR-α), which regulates the expression of genes that modulate fatty acid metabolism, can be regulated through interaction with coactivator and corepressor molecules. Understanding the nature of these interactions may help the development of drugs that effectively control lipid levels. By examining the crystal structure of a ternary complex comprised of the PPAR-α ligand-binding domain, an antagonist molecule, and the receptor-binding motif of the corepressor SMRT, Xu et al. report that a groove in the nuclear receptor contains overlapping binding sites for both coactivators and corepressors. However, in the presence of an antagonist molecule, a specific helix in the receptor is blocked from assuming an active conformation. This block results in a large domain that can specifically accommodate the receptor binding motif of a coprepressor. This mechanism may be utilized by nuclear receptors in general because the corepressor binding sequence is highly conserved. H. E. Xu, T. B. Stnaley, V. G. Montana, M. H. Lambert, B. G. Shearer, J. E. Cobb, D. D. McKee, C. M. Galardi, K. D. Plunket, R. T. Nolte, D. J. Parks, J. T. Moore, S. A. Kliewer, T. M.Willson, J. B. Stimmel, Structural basis for antagonist-mediated recruitment of nuclear co-repressors by PPARα. Nature 415 , 813-817 (2002). [Online Journal]

Structure of the PPARα and -γ Ligand Binding Domain in Complex with AZ 242; Ligand Selectivity and Agonist Activation in the PPAR Family

Background: The peroxisome proliferator-activated receptors (PPAR) are ligand-activated transcription factors belonging to the nuclear receptor family. The roles of PPARα in fatty acid oxidation and PPARγ in adipocyte differentiation and lipid storage have been characterized extensively. PPARs are activated by fatty acids and eicosanoids and are also targets for antidyslipidemic drugs, but the molecular interactions governing ligand selectivity for specific subtypes are unclear due to the lack of a PPARα ligand binding domain structure. Results: We have solved the crystal structure of the PPARα ligand binding domain (LBD) in complex with the combined PPARα and -γ agonist AZ 242, a novel dihydro cinnamate derivative that is structurally different from thiazolidinediones. In addition, we present the crystal structure of the PPARγ_LBD/AZ 242 complex and provide a rationale for ligand selectivity toward the PPARα and -γ subtypes. Heteronuclear NMR data on PPARα in both the apo form and in complex with AZ 242 shows an overall stabilization of the LBD upon agonist binding. A comparison of the novel PPARα/AZ 242 complex with the PPARγ/AZ 242 complex and previously solved PPARγ structures reveals a conserved hydrogen bonding network between agonists and the AF2 helix. Conclusions: The complex of PPARα and PPARγ with the dual specificity agonist AZ 242 highlights the conserved interactions required for receptor activation. Together with the NMR data, this suggests a general model for ligand activation in the PPAR family. A comparison of the ligand binding sites reveals a molecular explanation for subtype selectivity and provides a basis for rational drug design.

Pristanic acid and phytanic acid: naturally occurring ligands for the nuclear receptor peroxisome proliferator-activated receptor α

Phytanic acid and pristanic acid are branched-chain fatty acids, present at micromolar concentrations in the plasma of healthy individuals. Here we show that both phytanic acid and pristanic acid activate the peroxisome proliferator-activated receptor α (PPARα) in a concentration-dependent manner. Activation is observed via the ligand-binding domain of PPARα as well as via a PPAR response element (PPRE). Via the PPRE significant induction is found with both phytanic acid and pristanic acid at concentrations of 3 and 1 μM, respectively. The trans-activation of PPARδ and PPARγ by these two ligands is negligible. Besides PPARα, phytanic acid also trans-activates all three retinoic X receptor subtypes in a concentration-dependent manner. In primary human fibroblasts, deficient in phytanic acid α-oxidation, trans-activation through PPARα by phytanic acid is observed. This clearly demonstrates that phytanic acid itself, and not only its metabolite, pristanic acid, is a true physiological ligand for PPARα. Because induction of PPARα occurs at ligand concentrations comparable to the levels found for phytanic acid and pristanic acid in human plasma, these fatty acids should be seen as naturally occurring ligands for PPARα. These results demonstrate that both pristanic acid and phytanic acid are naturally occurring ligands for PPARα, which are present at physiological concentrations.—Zomer, A. W. M., B. van der Burg, G. A. Jansen, R. J. A. Wanders, B. T. Poll-The, and P. T. van der Saag. Pristanic acid and phytanic acid: naturally occurring ligands for the nuclear receptor peroxisome proliferator-activated receptor α. J. Lipid Res. 2000. 41: 1801–1807.

Role for peroxisome proliferator-activated receptor alpha in oxidized phospholipid-induced synthesis of monocyte chemotactic protein-1 and interleukin-8 by endothelial cells.

The attraction, binding, and entry of monocytes into the vessel wall play an important role in atherogenesis. We have previously shown that minimally oxidized/modified LDL (MM-LDL), a pathogenically relevant lipoprotein, can activate human aortic endothelial cells (HAECs) to produce monocyte chemotactic activators. In the present study, we demonstrate that MM-LDL and oxidation products of 1-palmitoyl-2-arachidonyl-sn-glycero-3-phosphocholine (PAPC) activate endothelial cells to synthesize monocyte chemotactic protein-1 (MCP-1) and interleukin-8 (IL-8). Several lines of evidence suggest that this activation is mediated by the lipid-dependent transcription factor peroxisome proliferator-activated receptor alpha (PPARalpha), the most abundant member of the PPAR family in HAECs. Treatment of transfected CV-1 cells demonstrated activation of the PPARalpha ligand-binding domain by MM-LDL, Ox-PAPC, or its component phospholipids, 1-palmitoyl-2-oxovalaroyl-sn-glycero-phosphocholine and 1-palmitoyl-2-glutaroyl-sn-glycero-phosphocholine; these lipids also activated a consensus peroxisome proliferator-activated receptor response element (PPRE) in transfected HAECs. Furthermore, activation of PPARalpha with synthetic ligand Wy14,643 stimulates the synthesis of IL-8 and MCP-1 by HAECs. By contrast, troglitazone, a PPARgamma agonist, decreased the levels of IL-8 and MCP-1. Finally, we demonstrate that unlike wild-type endothelial cells, endothelial cells derived from PPARalpha null mice do not produce MCP-1/JE in response to Ox-PAPC and MM-LDL. Together, these data demonstrate a proinflammatory role for PPARalpha in mediation of the activation of endothelial cells to produce monocyte chemotactic activity in response to oxidized phospholipids and lipoproteins.