The circadian clock acts at the genomic level to coordinate internal behavioral and physiologic rhythms via the CLOCK-BMAL transcriptional heterodimer. Although the nuclear receptors REV-ERBα and β have been proposed to form an accessory feedback loop that contributes to clock function1,2, their precise roles and importance remain unresolved. To establish their regulatory potential we generated comparative cistromes of both REV-ERB isoforms, which revealed shared recognition at over 50% of their total sites and extensive overlap with the master circadian regulator BMAL1. While Rev-erbα has been shown to directly regulate Bmal1 expression1,2, the cistromic analysis reveals a direct connection between Bmal1 and Rev-erbα and β regulatory circuits than previously suspected. Genes within the intersection of the BMAL1, REV-ERBα and REV-ERBβ cistromes are highly enriched for both clock and metabolic functions. As predicted by the cistromic analysis, dual depletion of Rev-erbα/β function by creating double-knockout mice (DKOs) profoundly disrupted circadian expression of core circadian clock and lipid homeostatic gene networks. As a result, DKOs show strikingly altered circadian wheel-running behavior and deregulated lipid metabolism. These data now ally Rev-erbα/β with Per, Cry and other components of the principal feedback loop that drives circadian expression and suggest a more integral mechanism for the coordination of circadian rhythm and metabolism.

Rev-Erbα and Rev-Erbβ are nuclear receptors that regulate the expression of genes involved in the control of circadian rhythm1,2, metabolism3,4, and inflammatory responses5. Rev-Erbs function as transcriptional repressors by recruiting NCoR/HDAC3 co-repressor complexes to Rev-Erb response elements in enhancers and promoters of target genes6-8, but the molecular basis for cell-specific programs of repression is not known. Here, we present evidence that in macrophages, Rev-Erbs regulate target gene expression by inhibiting the functions of distal enhancers that are selected by macrophage lineage-determining factors, thereby establishing a macrophage-specific program of repression. Remarkably, the repressive functions of Rev-Erbs are associated with their ability to inhibit the transcription of enhancer-derived RNAs (eRNAs). Furthermore, targeted degradation of eRNAs at two enhancers subject to negative regulation by Rev-Erbs resulted in reduced expression of nearby mRNAs, implying a direct role of these eRNAs in enhancer function. By precisely defining eRNA start sites using a method that quantifies nascent 5′ ends (5′-GRO-Seq), we show that transfer of full enhancer activity to a target promoter requires both the sequences mediating transcription factor binding and the specific sequences encoding the eRNA transcript. These studies provide evidence for direct roles of eRNAs in contributing to enhancer functions and suggest that Rev-Erbs act to suppress gene expression at a distance by repressing eRNA transcription.

Genome-wide studies have revealed that mammalian genomes are pervasively transcribed. This has led to the identification and isolation of novel classes of non-coding RNAs (ncRNAs) that influence gene expression by a variety of mechanisms. Here we review the characteristics and functions of regulatory ncRNAs in chromatin remodelling and at multiple levels of transcriptional and post-transcriptional regulation. We also describe the potential roles of ncRNAs in vascular biology and in mediating epigenetic modifications that might play roles in cardiovascular disease susceptibility. The emerging recognition of the diverse functions of ncRNAs in regulation of gene expression suggests that they may represent new targets for therapeutic intervention.

Macrophages play pivotal roles in both the induction and resolution phases of inflammatory processes. Macrophages have been shown to synthesize anti-inflammatory fatty acids in an LXR-dependent manner, but whether the production of these species contributes to the resolution phase of inflammatory responses has not been established. Here, we identify a biphasic program of gene expression that drives production of anti-inflammatory fatty acids 12–24h following TLR4 activation and contributes to down-regulation of mRNAs encoding pro-inflammatory mediators. Unexpectedly, rather than requiring LXRs, this late program of anti-inflammatory fatty acid biosynthesis is dependent on SREBP1 and results in the uncoupling of NFκB binding from gene activation. In contrast to previously identified roles of SREBP1 in promoting production of IL1β during the induction phase of inflammation, these studies provide evidence that SREBP1 also contributes to the resolution phase of TLR4-induced gene activation by reprogramming macrophage lipid metabolism.

Direct communication between arteries and veins without intervening capillary beds is the primary pathology of arteriovenous malformations (AVMs). Although Notch signaling is implicated in embryonic arteriovenous (AV) differentiation, its function in the adult mammalian vasculature has not been established due to the embryonic lethality that often occurs in both gain- and loss-of-function mutants. We expressed a constitutively active Notch4, int3, in the adult mouse endothelium by using the tetracycline-repressible system to suppress int3 during embryogenesis. int3 caused profound blood vessel enlargement and AV shunting, which are hallmarks of AVM, and led to lethality within weeks of its expression. Vessel enlargement, a manifestation of AVM, occurred in an apparently tissue-specific fashion; the liver, uterus, and skin were affected. int3-mediated vascular defects were accompanied by arterialization, including ectopic venous expression of ephrinB2, increased smooth muscle cells, and up-regulation of endogenous Notch signaling. Remarkably, the defective vessels and illness were reversed upon repression of int3 expression. Finally, endothelial expression of a constitutively active Notch1 induced similar hepatic vascular lesions. Our results provide gain-of-function evidence that Notch signaling in the adult endothelium is sufficient to render arterial characteristics and lead to AVMs

Brain arteriovenous malformations (BAVMs) can cause devastating stroke in young people and contribute to half of all hemorrhagic stroke in children. Unfortunately, the pathogenesis of BAVMs is unknown. In this article we show that activation of Notch signaling in the endothelium during brain development causes BAVM in mice. We turned on constitutively active Notch4 (int3) expression in endothelial cells from birth by using the tetracycline-regulatable system. All mutants developed hallmarks of BAVMs, including cerebral arteriovenous shunting and vessel enlargement, by 3 weeks of age and died by 5 weeks of age. Twenty-five percent of the mutants showed signs of neurological dysfunction, including ataxia and seizure. Affected mice exhibited hemorrhage and neuronal cell death within the cerebral cortex and cerebellum. Strikingly, int3 repression resolved ataxia and reversed the disease progression, demonstrating that int3 is not only sufficient to induce, but also required to sustain the disease. We show that int3 expression results in widespread enlargement of the microvasculature, which coincided with a reduction in capillary density, linking vessel enlargement to Notch's known function of inhibiting vessel sprouting. Our data suggest that the Notch pathway is a molecular regulator of BAVM pathogenesis in mice, and offer hope that their regression might be possible by targeting the causal molecular lesion.angiogenesis ͉ cell signaling ͉ endothelial cell ͉ stroke ͉ cerebrovascular

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Hemophagocytic lymphohistiocytosis (HLH) is characterized by immune dysregulation due to inadequate restraint of overactivated immune cells and is associated with a variable clinical spectrum having overlap with more common pathophysiologies. HLH is difficult to diagnose and can be part of inflammatory syndromes. Here, we identify a novel hematological/autoinflammatory condition (NOCARH syndrome) in four unrelated patients with superimposable features, including neonatal-onset cytopenia with dyshematopoiesis, autoinflammation, rash, and HLH. Patients shared the same de novo CDC42 mutation (Chr1:22417990CT, p.R186C) and altered hematopoietic compartment, immune dysregulation, and inflammation. CDC42 mutations had been associated with syndromic neurodevelopmental disorders. In vitro and in vivo assays documented unique effects of p.R186C on CDC42 localization and function, correlating with the distinctiveness of the trait. Emapalumab was critical to the survival of one patient, who underwent successful bone marrow transplantation. Early recognition of the disorder and establishment of treatment followed by bone marrow transplant are important to survival.