Grant D. Barish

PPARδ: a dagger in the heart of the metabolic syndrome

Obesity is a growing threat to global health by virtue of its association with insulin resistance, glucose intolerance, hypertension, and dyslipidemia, collectively known as the metabolic syndrome or syndrome X. The nuclear receptors PPARα and PPARγ are therapeutic targets for hypertriglyceridemia and insulin resistance, respectively, and drugs that modulate these receptors are currently in clinical use. More recent work on the less-described PPAR isotype PPARδ has uncovered a dual benefit for both hypertriglyceridemia and insulin resistance, highlighting the broad potential of PPARδ in the treatment of metabolic disease. PPARδ enhances fatty acid catabolism and energy uncoupling in adipose tissue and muscle, and it suppresses macrophage-derived inflammation. Its combined activities in these and other tissues make it a multifaceted therapeutic target for the metabolic syndrome with the potential to control weight gain, enhance physical endurance, improve insulin sensitivity, and ameliorate atherosclerosis.

Regulation of circadian behaviour and metabolism by REV-ERB-α and REV-ERB-β

The circadian clock acts at the genomic level to coordinate internal behavioural and physiological rhythms via the CLOCK–BMAL1 transcriptional heterodimer. Although the nuclear receptors REV-ERB-α and REV-ERB-β have been proposed to form an accessory feedback loop that contributes to clock function, their precise roles and importance remain unresolved. To establish their regulatory potential, we determined the genome-wide cis-acting targets (cistromes) of both REV-ERB isoforms in murine liver, which revealed shared recognition at over 50% of their total DNA binding sites and extensive overlap with the master circadian regulator BMAL1. Although REV-ERB-α has been shown to regulate Bmal1 expression directly, our cistromic analysis reveals a more profound connection between BMAL1 and the REV-ERB-α and REV-ERB-β genomic regulatory circuits than was previously suspected. Genes within the intersection of the BMAL1, REV-ERB-α and REV-ERB-β cistromes are highly enriched for both clock and metabolic functions. As predicted by the cistromic analysis, dual depletion of Rev-erb-α and Rev-erb-β function by creating double-knockout mice profoundly disrupted circadian expression of core circadian clock and lipid homeostatic gene networks. As a result, double-knockout mice show markedly altered circadian wheel-running behaviour and deregulated lipid metabolism. These data now unite REV-ERB-α and REV-ERB-β with PER, CRY and other components of the principal feedback loop that drives circadian expression and indicate a more integral mechanism for the coordination of circadian rhythm and metabolism.

Regulation of Wnt Signaling by Sox Proteins: XSox17α/β and XSox3 Physically Interact with β-catenin

Abstract Using a functional screen in Xenopus embryos, we identified a novel function for the HMG box protein XSox17β. Ectopic expression of XSox17β ventralizes embryos by inhibiting the Wnt pathway downstream of β-catenin but upstream of the Wnt-responsive gene Siamois . XSox17β also represses transactivation of a TCF/LEF-dependent reporter construct by Wnt and β-catenin. In animal cap experiments, it both activates transcription of endodermal genes and represses β-catenin-stimulated expression of dorsal genes. The inhibition activity of XSox17β maps to a region C-terminal to the HMG box; this region of XSox17β physically interacts with the Armadillo repeats of β-catenin. Two additional Sox proteins, XSox17α and XSox3, likewise bind to β-catenin and inhibit its TCF-mediated signaling activity. These results reveal an unexpected mechanism by which Sox proteins can modulate Wnt signaling pathways.

A Vitamin D Receptor/SMAD Genomic Circuit Gates Hepatic Fibrotic Response

Liver fibrosis is a reversible wound-healing response involving TGFβ1/SMAD activation of hepatic stellate cells (HSCs). It results from excessive deposition of extracellular matrix components and can lead to impairment of liver function. Here, we show that vitamin D receptor (VDR) ligands inhibit HSC activation by TGFβ1 and abrogate liver fibrosis, whereas Vdr knockout mice spontaneously develop hepatic fibrosis. Mechanistically, we show that TGFβ1 signaling causes a redistribution of genome-wide VDR-binding sites (VDR cistrome) in HSCs and facilitates VDR binding at SMAD3 profibrotic target genes via TGFβ1-dependent chromatin remodeling. In the presence of VDR ligands, VDR binding to the coregulated genes reduces SMAD3 occupancy at these sites, inhibiting fibrosis. These results reveal an intersecting VDR/SMAD genomic circuit that regulates hepatic fibrogenesis and define a role for VDR as an endocrine checkpoint to modulate the wound-healing response in liver. Furthermore, the findings suggest VDR ligands as a potential therapy for liver fibrosis.

Cryptochromes mediate rhythmic repression of the glucocorticoid receptor

Mammalian metabolism is highly circadian and major hormonal circuits involving nuclear hormone receptors display interlinked diurnal cycling. However, mechanisms that logically explain the coordination of nuclear hormone receptors and the clock are poorly understood. Here we show that two circadian co-regulators, cryptochromes 1 and 2, interact with the glucocorticoid receptor in a ligand-dependent fashion and globally alter the transcriptional response to glucocorticoids in mouse embryonic fibroblasts: cryptochrome deficiency vastly decreases gene repression and approximately doubles the number of dexamethasone-induced genes, suggesting that cryptochromes broadly oppose glucocorticoid receptor activation and promote repression. In mice, genetic loss of cryptochrome 1 and/or 2 results in glucose intolerance and constitutively high levels of circulating corticosterone, suggesting reduced suppression of the hypothalamic–pituitary–adrenal axis coupled with increased glucocorticoid transactivation in the liver. Genomically, cryptochromes 1 and 2 associate with a glucocorticoid response element in the phosphoenolpyruvate carboxykinase 1 promoter in a hormone-dependent manner, and dexamethasone-induced transcription of the phosphoenolpyruvate carboxykinase 1 gene was strikingly increased in cryptochrome-deficient livers. These results reveal a specific mechanism through which cryptochromes couple the activity of clock and receptor target genes to complex genomic circuits underpinning normal metabolic homeostasis.

PPARδ-mediated antiinflammatory mechanisms inhibit angiotensin II-accelerated atherosclerosis

Activation of the nuclear hormone receptor peroxisome proliferator-activated receptor δ (PPARδ) has been shown to improve insulin resistance, adiposity, and plasma HDL levels. However, its antiatherogenic role remains controversial. Here we report atheroprotective effects of PPARδ activation in a model of angiotensin II (AngII)-accelerated atherosclerosis, characterized by increased vascular inflammation related to repression of an antiinflammatory corepressor, B cell lymphoma-6 (Bcl-6), and the regulators of G protein-coupled signaling (RGS) proteins RGS4 and RGS5. In this model, administration of the PPARδ agonist GW0742 (1 or 10 mg/kg) substantially attenuated AngII-accelerated atherosclerosis without altering blood pressure and increased vascular expression of Bcl-6, RGS4, and RGS5, which was associated with suppression of inflammatory and atherogenic gene expression in the artery. In vitro studies demonstrated similar changes in AngII-treated macrophages: PPARδ activation increased both total and free Bcl-6 levels and inhibited AngII activation of MAP kinases, p38, and ERK1/2. These studies uncover crucial proinflammatory mechanisms of AngII and highlight actions of PPARδ activation to inhibit AngII signaling, which is atheroprotective.

Bcl-6 and NF-κB cistromes mediate opposing regulation of the innate immune response

In the macrophage, toll-like receptors (TLRs) are key sensors that trigger signaling cascades to activate inflammatory programs via the NF-κB gene network. However, the genomic network targeted by TLR/NF-κB activation and the molecular basis by which it is restrained to terminate activation and re-establish quiescence is poorly understood. Here, using chromatin immunoprecipitation sequencing (ChIP-seq), we define the NF-κB cistrome, which is comprised of 31,070 cis-acting binding sites responsive to lipopolysaccharide (LPS)-induced signaling. In addition, we demonstrate that the transcriptional repressor B-cell lymphoma 6 (Bcl-6) regulates nearly a third of the Tlr4-regulated transcriptome, and that 90% of the Bcl-6 cistrome is collapsed following Tlr4 activation. Bcl-6-deficient macrophages are acutely hypersensitive to LPS and, using comparative ChIP-seq analyses, we found that the Bcl-6 and NF-κB cistromes intersect, within nucleosomal distance, at nearly half of Bcl-6-binding sites in stimulated macrophages to promote opposing epigenetic modifications of the local chromatin. These results reveal a genomic strategy for controlling the innate immune response in which repressive and inductive cistromes establish a dynamic balance between macrophage quiescence and activation via epigenetically marked cis-regulatory elements.

PPARγ activation in adipocytes is sufficient for systemic insulin sensitization

Although peroxisome proliferator-activated receptor gamma (PPARγ) agonists such as thiazolidinediones (TZDs) are widely used to treat type 2 diabetes, how its activation in individual tissues contributes to TZDs therapeutic action remains controversial. As TZDs are known to have receptor-independent effects, we sought to establish gain-of-function animal models to delineate the receptors insulin-sensitizing actions. Unexpectedly, we find that selective activation of PPARγ in adipocytes, but not in macrophages, is sufficient for whole-body insulin sensitization equivalent to systemic TZD treatment. In addition to improved adipokine, inflammatory, and lipid profiles, PPARγ activation in mature adipocytes normalizes serum insulin without increased adipogenesis. Co-culture studies indicated that PPARγ-activated adipocytes broadly suppress induction of inflammatory cytokines and C-X-C family chemokines in macrophages. Collectively, these data describe an “adipocentric” model in which adipose activation of PPARγ is sufficient for complete insulin sensitization and suggest a specific application for fat selective PPARγ modulators in diabetic therapy.

PPARδ regulates multiple proinflammatory pathways to suppress atherosclerosis

Lipid homeostasis and inflammation are key determinants in atherogenesis, exemplified by the requirement of lipid-laden, foam cell macrophages for atherosclerotic lesion formation. Although the nuclear receptor PPARδ has been implicated in both systemic lipid metabolism and macrophage inflammation, its role as a therapeutic target in vascular disease is unclear. We show here that orally active PPARδ agonists significantly reduce atherosclerosis in apoE−/− mice. Metabolic and gene expression studies reveal that PPARδ attenuates lesion progression through its HDL-raising effect and anti-inflammatory activity within the vessel wall, where it suppresses chemoattractant signaling by down-regulation of chemokines. Activation of PPARδ also induces the expression of regulator of G protein signaling (RGS) genes, which are implicated in blocking the signal transduction of chemokine receptors. Consistent with this, PPARδ ligands repress monocyte transmigration and macrophage inflammatory responses elicited by atherogenic cytokines. These results reveal that PPARδ antagonizes multiple proinflammatory pathways and suggest PPARδ-selective drugs as candidate therapeutics for atherosclerosis.

NCoR1 is a conserved physiological modulator of muscle mass and oxidative function.

Transcriptional coregulators control the activity of many transcription factors and are thought to have wide-ranging effects on gene expression patterns. We show here that muscle-specific loss of nuclear receptor corepressor 1 (NCoR1) in mice leads to enhanced exercise endurance due to an increase of both muscle mass and of mitochondrial number and activity. The activation of selected transcription factors that control muscle function, such as MEF2, PPARβ/δ, and ERRs, underpins these phenotypic alterations. NCoR1 levels are decreased in conditions that require fat oxidation, resetting transcriptional programs to boost oxidative metabolism. Knockdown of gei-8, the sole C. elegans NCoR homolog, also robustly increased muscle mitochondria and respiration, suggesting conservation of NCoR1 function. Collectively, our data suggest that NCoR1 plays an adaptive role in muscle physiology and that interference with NCoR1 action could be used to improve muscle function.