Identification of the fructose transporter GLUT5 (SLC2A5) as a novel target of nuclear receptor LXR

Fructose has become a major constituent of our modern diet and is implicated as an underlying cause in the development of metabolic diseases. The fructose transporter GLUT5 (SLC2A5) is required for intestinal fructose absorption. GLUT5 expression is induced in the intestine and skeletal muscle of type 2 diabetes (T2D) patients and in certain cancers that are dependent on fructose metabolism, indicating that modulation of GLUT5 levels could have potential in the treatment of these diseases. Using an unbiased screen for transcriptional control of the human GLUT5 promoter we identified a strong and specific regulation by liver X receptor alpha (LXR alpha, NR1H3). Using promoter truncations and site-directed mutagenesis we identified a functional LXR response element (LXRE) in the human GLUT5 promoter, located at -385 bp relative to the transcriptional start site (TSS). Finally, mice treated with LXR agonist T0901317 showed an increase in Glut5 mRNA and protein levels in duodenum and adipose tissue, underscoring the in vivo relevance of its regulation by LXR. Together, our findings show that LXR alpha regulates GLUT5 in mice and humans. As a ligand-activated transcription factor, LXR alpha might provide novel pharmacologic strategies for the selective modulation of GLUT5 activity in the treatment of metabolic disease as well as cancer.</p

MRF4 negatively regulates adult skeletal muscle growth by repressing MEF2 activity

The myogenic regulatory factor MRF4 is highly expressed in adult skeletal muscle but its function is unknown. Here we show that Mrf4 knockdown in adult muscle induces hypertrophy and prevents denervation-induced atrophy. This effect is accompanied by increased protein synthesis and widespread activation of muscle-specific genes, many of which are targets of MEF2 transcription factors. MEF2-dependent genes represent the top-ranking gene set enriched after Mrf4 RNAi and a MEF2 reporter is inhibited by co-transfected MRF4 and activated by Mrf4 RNAi. The Mrf4 RNAi-dependent increase in fibre size is prevented by dominant negative MEF2, while constitutively active MEF2 is able to induce myofibre hypertrophy. The nuclear localization of the MEF2 corepressor HDAC4 is impaired by Mrf4 knockdown, suggesting that MRF4 acts by stabilizing a repressor complex that controls MEF2 activity. These findings open new perspectives in the search for therapeutic targets to prevent muscle wasting, in particular sarcopenia and cachexia

Machine learning reveals STAT motifs as predictors for GR-mediated gene repression

Glucocorticoids are potent immunosuppressive drugs, but long-term treatment leads to severe side-effects. While there is a commonly accepted model for GR-mediated gene activation, the mechanism behind repression remains elusive. Understanding the molecular action of the glucocorticoid receptor (GR) mediated gene repression is the first step towards developing novel therapies. We devised an approach that combines multiple epigenetic assays with 3D chromatin data to find sequence patterns predicting gene expression change. We systematically tested> 100 models to evaluate the best way to integrate the data types and found that GR-bound regions hold most of the information needed to predict the polarity of Dex-induced transcriptional changes. We confirmed NF-κB motif family members as predictors for gene repression and identified STAT motifs as additional negative predictors

FRIGIDA-Independent Variation in Flowering Time of Natural Arabidopsis thaliana Accessions

FRIGIDA (FRI) and FLOWERING LOCUS C (FLC) are two genes that, unless plants are vernalized, greatly delay flowering time in Arabidopsis thaliana. Natural loss-of-function mutations in FRI cause the early flowering growth habits of many A. thaliana accessions. To quantify the variation among wild accessions due to FRI, and to identify additional genetic loci in wild accessions that influence flowering time, we surveyed the flowering times of 145 accessions in long-day photoperiods, with and without a 30-day vernalization treatment, and genotyped them for two common natural lesions in FRI. FRI is disrupted in at least 84 of the accessions, accounting for only ∼40% of the flowering-time variation in long days. During efforts to dissect the causes for variation that are independent of known dysfunctional FRI alleles, we found new loss-of-function alleles in FLC, as well as late-flowering alleles that do not map to FRI or FLC. An FLC nonsense mutation was found in the early flowering Van-0 accession, which has otherwise functional FRI. In contrast, Lz-0 flowers late because of high levels of FLC expression, even though it has a deletion in FRI. Finally, eXtreme array mapping identified genomic regions linked to the vernalization-independent, late-flowering habit of Bur-0, which has an alternatively spliced FLC allele that behaves as a null allele

Dissection of floral induction pathways using global expression analysis

Flowering of the reference plant Arabidopsis thaliana is controlled by several signaling pathways, which converge on a small set of genes that function as pathway integrators. We have analyzed the genomic response to one type of floral inductive signal, photoperiod, to dissect the function of several genes transducing this stimulus, including CONSTANS, thought to be the major output of the photoperiod pathway. Comparing the effects of CONSTANS with those of FLOWERING LOCUS T, which integrates inputs from CONSTANS and other floral inductive pathways, we find that expression profiles of shoot apices from plants with mutations in either gene are very similar. In contrast, a mutation in LEAFY, which also acts downstream of CONSTANS, has much more limited effects. Another pathway integrator, SUPPRESSOR OF OVEREXPRESSION OF CO 1, is responsive to acute induction by photoperiod even in the presence of the floral repressor encoded by FLOWERING LOCUS C. We have discovered a large group of potential floral repressors that are down-regulated upon photoperiodic induction. These include two AP2 domain-encoding genes that can repress flowering. The two paralogous genes, SCHLAFMUTZE and SCHNARCHZAPFEN, share a signature with partial complementarity to the miR172 microRNA, whose precursor we show to be induced upon flowering. These and related findings on SPL genes suggest that microRNAs play an important role in the regulation of flowering

E47 modulates hepatic glucocorticoid action

Glucocorticoids (GCs) are effective drugs, but their clinical use is compromised by severe side effects including hyperglycemia, hyperlipidemia and obesity. They bind to the Glucocorticoid Receptor (GR), which acts as a transcription factor. The activation of metabolic genes by GR is thought to underlie these adverse effects. We identify the bHLH factor E47 as a modulator of GR target genes. Using mouse genetics, we find that E47 is required for the regulation of hepatic glucose and lipid metabolism by GR, and that loss of E47 prevents the development of hyperglycemia and hepatic steatosis in response to GCs. Here we show that E47 and GR co-occupy metabolic promoters and enhancers. E47 is needed for the efficient recruitment of GR and coregulators such as Mediator to chromatin. Altogether, our results illustrate how GR and E47 regulate hepatic metabolism, and might provide an entry point for novel therapies with reduced side effects

Rapid Genome-wide Recruitment of RNA Polymerase II Drives Transcription, Splicing, and Translation Events during T Cell Responses

Summary: Activation of immune cells results in rapid functional changes, but how such fast changes are accomplished remains enigmatic. By combining time courses of 4sU-seq, RNA-seq, ribosome profiling (RP), and RNA polymerase II (RNA Pol II) ChIP-seq during T cell activation, we illustrate genome-wide temporal dynamics for ∼10,000 genes. This approach reveals not only immediate-early and posttranscriptionally regulated genes but also coupled changes in transcription and translation for >90% of genes. Recruitment, rather than release of paused RNA Pol II, primarily mediates transcriptional changes. This coincides with a genome-wide temporary slowdown in cotranscriptional splicing, even for polyadenylated mRNAs that are localized at the chromatin. Subsequent splicing optimization correlates with increasing Ser-2 phosphorylation of the RNA Pol II carboxy-terminal domain (CTD) and activation of the positive transcription elongation factor (pTEFb). Thus, rapid de novo recruitment of RNA Pol II dictates the course of events during T cell activation, particularly transcription, splicing, and consequently translation. : Davari et al. visualize global changes in RNA Pol II binding, transcription, splicing, and translation. T cells change their functional program by rapid de novo recruitment of RNA Pol II and coupled changes in transcription and translation. This coincides with fluctuations in RNA Pol II phosphorylation and a temporary reduction in cotranscriptional splicing. Keywords: RNA Pol II, cotranscriptional splicing, T cell activation, ribosome profiling, 4sU, H3K36, Ser-5 RNA Pol II, Ser-2 RNA Pol II, immune response, immediate-early gene

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(1,2). 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

Dicer in Macrophages Prevents Atherosclerosis by Promoting Mitochondrial Oxidative Metabolism

Background: Alternative macrophage activation, which relies on mitochondrial oxidative metabolism, plays a central role in the resolution of inflammation and prevents atherosclerosis. Moreover, macrophages handle large amounts of cholesterol and triglycerides derived from the engulfed modified lipoproteins during atherosclerosis. Although several microRNAs regulate macrophage polarization, the role of the microRNA-generating enzyme Dicer in macrophage activation during atherosclerosis is unknown. Methods: To evaluate the role of Dicer in atherosclerosis, Apoe(-/-) mice with or without macrophage-specific Dicer deletion were fed a high-fat diet for 12 weeks. Anti-argonaute 2 RNA immunoprecipitation chip and RNA deep sequencing combined with microRNA functional screening were performed in the Dicer wild-type and knockout bone marrow-derived macrophages to identify the individual microRNAs and the mRNA targets mediating the phenotypic effects of Dicer. The role of the identified individual microRNA and its target in atherosclerosis was determined by tail vein injection of the target site blockers in atherosclerotic Apoe(-/-) mice. Results: We show that Dicer deletion in macrophages accelerated atherosclerosis in mice, along with enhanced inflammatory response and increased lipid accumulation in lesional macrophages. In vitro, alternative activation was limited whereas lipid-filled foam cell formation was exacerbated in Dicer-deficient macrophages as a result of impaired mitochondrial fatty acid oxidative metabolism. Rescue of microRNA (miR)-10a, let-7b, and miR-195a expression restored the oxidative metabolism in alternatively activated Dicer-deficient macrophages. Suppression of ligand-dependent nuclear receptor corepressor by miR-10a promoted fatty acid oxidation, which mediated the lipolytic and anti-inflammatory effect of Dicer. miR-10a expression was negatively correlated to the progression of atherosclerosis in humans. Blocking the interaction between ligand-dependent nuclear receptor corepressor and miR-10a by target site blockers aggravated atherosclerosis development in mice. Conclusions: Dicer plays an atheroprotective role by coordinately regulating the inflammatory response and lipid metabolism in macrophages through enhancing fatty acid-fueled mitochondrial respiration, suggesting that promoting Dicer/miR-10a-dependent metabolic reprogramming in macrophages has potential therapeutic implications to prevent atherosclerosis