Metabolites: a helping hand for pathway evolution?

The evolution of enzymes and pathways is under debate. Recent studies show that recruitment of single enzymes from different pathways could be the driving force for pathway evolution. Other mechanisms of evolution, such as pathway duplication, enzyme specialization, de novo invention of pathways or retro-evolution of pathways, appear to be less abundant. Twenty percent of enzyme superfamilies are quite variable, not only in changing reaction chemistry or metabolite type but in changing both at the same time. These variable superfamilies account for nearly half of all known reactions. The most frequently occurring metabolites provide a helping hand for such changes because they can be accommodated by many enzyme superfamilies. Thus, a picture is emerging in which new pathways are evolving from central metabolites by preference, thereby keeping the overall topology of the metabolic network.     

PPARγ agonists induce a white-to-brown fat conversion through stabilization of PRDM16 protein

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Highlights? PPARγ full agonists induce a brown fat phenotype in subcutaneous WAT ? PRDM16 is required for the development of PPARγ agonist-inducible brow adipocytes ? PRDM16 and PPARγ agonists synergistically activate the brown fat gene program ? Browning effect is mediated through the enhanced stability of the PRDM16 protein     

Exercise-associated generation of PPARγ ligands activates PPARγ signaling events and upregulates genes related to lipid metabolism

The aim of the present study was to test the hypotheses that exercise is associated with generation of peroxisome proliferator-activated receptor-γ (PPARγ) ligands in the plasma and that this may activate PPARγ signaling within circulating monocytes, thus providing a mechanism to underpin the exercise-induced antiatherogenic benefits observed in previous studies. A cohort of healthy individuals undertook an 8-wk exercise-training program; samples were obtained before (Pre) and after (Post) standardized submaximal exercise bouts (45 min of cycling at 70% of maximal O(2) uptake, determined at baseline) at weeks 0, 4, and 8. Addition of plasma samples to PPARγ response element (PPRE)-luciferase reporter gene assays showed increased PPARγ activity following standardized exercise bouts (Post/Pre = 1.23 ± 0.10 at week 0, P < 0.05), suggesting that PPARγ ligands were generated during exercise. However, increases in PPARγ/PPRE-luciferase activity in response to the same standardized exercise bout were blunted during the training program (Post/Pre = 1.18 ± 0.14 and 1.10 ± 0.10 at weeks 4 and 8, respectively, P > 0.05 for both), suggesting that the relative intensity of the exercise may affect PPARγ ligand generation. In untrained individuals, specific transient increases in monocyte expression of PPARγ-regulated genes were observed within 1.5-3 h of exercise (1.7 ± 0.4, 2.6 ± 0.4, and 1.4 ± 0.1 fold for CD36, liver X receptor-α, and ATP-binding cassette subfamily A member 1, respectively, P < 0.05), with expression returning to basal levels within 24 h. In contrast, by the end of the exercise program, expression at the protein level of PPARγ target genes had undergone sustained increases that were not associated with an individual exercise bout (e.g., week 8 Pre/week 0 Pre = 2.79 ± 0.61 for CD36, P < 0.05). Exercise is known to upregulate PPARγ-controlled genes to induce beneficial effects in skeletal muscle (e.g., mitochondrial biogenesis and aerobic respiration). We suggest that parallel exercise-induced benefits may occur in monocytes, as monocyte PPARγ activation has been linked to beneficial antidiabetic effects (e.g., exercise-induced upregulation of monocytic PPARγ-controlled genes is associated with reverse cholesterol transport and anti-inflammatory effects). Thus, exercise-triggered monocyte PPARγ activation may constitute an additional rationale for prescribing exercise to type 2 diabetes patients.     

Revealing a steroid receptor ligand as a unique PPARγ agonist

Peroxisome proliferator-activated receptor gamma (PPARγ) regulates metabolic homeostasis and is a molecular target for anti-diabetic drugs. We report here the identification of a steroid receptor ligand, RU-486, as an unexpected PPARγ agonist, thereby uncovering a novel signaling route for this steroid drug. Similar to rosiglitazone, RU-486 modulates the expression of key PPARγ target genes and promotes adipocyte differentiation, but with a lower adipogenic activity. Structural and functional studies of receptor-ligand interactions reveal the molecular basis for a unique binding mode for RU-486 in the PPARγ ligand-binding pocket with distinctive properties and epitopes, providing the molecular mechanisms for the discrimination of RU-486 from thiazolidinediones (TZDs) drugs. Our findings together indicate that steroid compounds may represent an alternative approach for designing non-TZD PPARγ ligands in the treatment of insulin resistance.     

Structure optimization of a new class of PPARγ antagonists

Peroxisome proliferator-activated receptor gamma (PPARγ) modulators have found wide application for the treatment of cancers, metabolic disorders and inflammatory diseases. Contrary to PPARγ agonists, PPARγ antagonists have been much less studied and although they have shown immunomodulatory effects, there is still no therapeutically useful PPARγ antagonist on the market. In contrast to non-competitive, irreversible inhibition caused by 2-chloro-5-nitrobenzanilide (GW9662), the recently described (E)-2-(5-((4-methoxy-2-(trifluoromethyl)quinolin-6-yl)methoxy)-2-((4-(trifluoromethyl)benzyl)oxy)-benzylidene)-hexanoic acid (MTTB, T-10017) is a promising prototype for a new class of PPARγ antagonists. It exhibits competitive antagonism against rosiglitazone mediated activation of PPARγ ligand binding domain (PPARγLBD) in a transactivation assay in HEK293T cells with an IC₅₀ of 4.3 µM against 1 µM rosiglitazone. The aim of this study was to investigate the structure-activity relationships (SAR) of the MTTB scaffold focusing on improving its physicochemical properties. Through this optimization, 34 new derivatives were prepared and characterized. Two new potent compounds (T-10075 and T-10106) with much improved drug-like properties and promising pharmacokinetic profile were identified.     

X-ray crystal structure of rivoglitazone bound to PPARγ and PPAR subtype selectivity of TZDs

Thiazolidinedione (TZD) compounds targeting the nuclear receptor peroxisome proliferator-activated receptor gamma (PPARγ) demonstrate unique benefits for the treatment of insulin resistance and type II diabetes. TZDs include rosiglitazone, pioglitazone and rivoglitazone, with the latter being the most potent. The TZDs are only marginally selective for the therapeutic target PPARγ as they also activate PPARα and PPARδ homologues to varying degrees, causing off-target effects. While crystal structures for TZD compounds in complex with PPARγ are available, minimal structural information is available for TZDs bound to PPARα and PPARδ. This paucity of structural information has hampered the determination of precise structural mechanisms involved in TZD selectivity between PPARs. To help address these questions molecular dynamic simulations were performed of rosiglitazone, pioglitazone and rivoglitazone in complex with PPARα, PPARδ, and PPARγ in order to better understand the mechanisms of PPAR selectivity. The simulations revealed that TZD interactions with residues Tyr314 and Phe318 of PPARα and residues Phe291 and Thr253 of PPARδ as well as the omega loop, are key determinants of TZD receptor selectivity. Notably, in this study, we solve the first X-ray crystal structure of rivoglitazone bound to any PPAR. Rivoglitazone forms a unique hydrogen bond network with the residues of the PPARγ co-activator binding surface (known as AF2) and makes more extensive contacts with helix 3 and the β-sheet as compared to model TZD compounds such as rosiglitazone.     

5-Lipoxygenase contributes to PPARγ activation in macrophages in response to apoptotic cells

Macrophage polarization to an anti-inflammatory phenotype upon contact with apoptotic cells is a contributing hallmark to immune suppression during the late phase of sepsis. Although the peroxisome proliferator-activated receptor γ (PPARγ) supports this macrophage phenotype switch, it remains elusive how apoptotic cells activate PPARγ. Assuming that a molecule causing PPARγ activation in macrophages originates in the cell membrane of apoptotic cells we analyzed lipid rafts from apoptotic, necrotic, and living human Jurkat T cells which showed the presence of 5-lipoxygenase (5-LO) in lipid rafts of apoptotic cells only. Incubating macrophages with lipid rafts of apoptotic, but not necrotic or living cells, induced PPAR responsive element (PPRE)-driven mRuby reporter gene expression in RAW 264.7 macrophages stably transduced with a 4xPPRE containing vector. Experiments with lipid rafts of apoptotic murine EL4 T cells revealed similar results. To verify the involvement of 5-LO in activating PPARγ in macrophages, Jurkat T cells were incubated with the 5-LO inhibitor MK-866 prior to induction of apoptosis, which failed to induce mRuby expression. Similar results were obtained with lipid rafts of apoptotic EL4 T cells preexposed to the 5-LO inhibitors zileuton and CJ-13610. Interestingly, Jurkat T cells overexpressing 5-LO failed to activate PPARγ in macrophages, while their 5-LO overexpressing apoptotic counterparts did. Our results suggest that during apoptosis 5-LO gets associated with lipid rafts and synthesizes ligands that in turn stimulate PPARγ in macrophages.     

Adipocyte NCoR knockout decreases PPARγ phosphorylation and enhances PPARγ activity and insulin sensitivity

Insulin resistance, tissue inflammation, and adipose tissue dysfunction are features of obesity and Type 2 diabetes. We generated adipocyte-specific Nuclear Receptor Corepressor (NCoR) knockout (AKO) mice to investigate the function of NCoR in adipocyte biology, glucose and insulin homeostasis. Despite increased obesity, glucose tolerance was improved in AKO mice, and clamp studies demonstrated enhanced insulin sensitivity in liver, muscle, and fat. Adipose tissue macrophage infiltration and inflammation were also decreased. PPARγ response genes were upregulated in adipose tissue from AKO mice and CDK5-mediated PPARγ ser-273 phosphorylation was reduced, creating a constitutively active PPARγ state. This identifies NCoR as an adaptor protein that enhances the ability of CDK5 to associate with and phosphorylate PPARγ. The dominant function of adipocyte NCoR is to transrepress PPARγ and promote PPARγ ser-273 phosphorylation, such that NCoR deletion leads to adipogenesis, reduced inflammation, and enhanced systemic insulin sensitivity, phenocopying the TZD-treated state.     

Eye-hand coordination: eye to hand or hand to eye?

Single-unit recording has revealed both hand and eye movement-related activity in the parietal cortex of the macaque monkey. These experiments, as well as neuropsychological studies, are unravelling the complex nature of how the eye and the hand work together in the control of visually guided movements.     

Revisiting PPARγ as a target for the treatment of metabolic disorders

As the prevalence of obesity has increased explosively over the last several decades, associated metabolic disorders, including type 2 diabetes, dyslipidemia, hypertension, and cardiovascular diseases, have been also increased. Thus, new strategies for preventing and treating them are needed. The nuclear peroxisome proliferator-activated receptors (PPARs) are involved fundamentally in regulating energy homeostasis; thus, they have been considered attractive drug targets for addressing metabolic disorders. Among the PPARs, PPARγ is a master regulator of gene expression for metabolism, inflammation, and other pathways in many cell types, especially adipocytes. It is a physiological receptor of the potent anti-diabetic drugs of the thiazolidinediones (TZDs) class, including rosiglitazone (Avandia). However, TZDs have undesirable and severe side effects, such as weight gain, fluid retention, and cardiovascular dysfunction. Recently, many reports have suggested that PPARγ could be modulated by post-translational modifications (PTMs), and modulation of PTM has been considered as novel approaches for treating metabolic disorders with fewer side effects than the TZDs. In this review, we discuss how PTM of PPARγ may be regulated and issues to be considered in making novel anti-diabetic drugs that can modulate the PTM of PPARγ. [BMB Reports 2014; 47(11): 599-608]     

Discovery of a novel selective PPARγ modulator from (−)-Cercosporamide derivatives

In an investigation of (?)-Cercosporamide derivatives with a plasma glucose-lowering effect, we found that N-benzylcarboxamide derivative 4 was a partial agonist of PPARγ. A SAR study of the substituents on carboxamide nitrogen afforded the N-(1-naphthyl)methylcarboxamide derivative 23 as the most potent selective PPARγ modulator. An X-ray crystallography study revealed that compound 23 bounded to the PPARγ ligand binding domain in a unique way without any interaction with helix12. Compound 23 displayed a potent plasma glucose-lowering effect in db/db mice without the undesirable increase in body fluid and heart weight that is typically observed when PPARγ full agonists are administrated.     

Development of a new class of benzoylpyrrole-based PPARα/γ activators

Starting with a subtle blood glucose-lowering effect of a TGF-β inhibitor, we designed and synthesized a series of benzoylpyrrole-based carboxylic acids as PPARs activators. Among these compounds, 10sNa exhibited favorable blood glucose-lowering effect without body weight gain. We assume that the beneficial effect of 10sNa is attributed to not only its compound PPARα agonistic activity but also its PPARγ partial agonistic activity.