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Biology subjects

Ray, D. W.

Publications and source records attributed to Ray, D. W..

10 recordsLinked to original sources

Fibroblast-like synoviocytes orchestrate daily rhythmic inflammation in arthritis

Rheumatoid arthritis is a chronic inflammatory disease that shows characteristic diurnal variation in symptom severity, where joint resident fibroblast-like synoviocytes (FLS) act as important mediators of arthritis pathology. We investigate the role of FLS circadian clock function in directing rhythmic joint inflammation in a murine model of inflammatory arthritis. We demonstrate FLS time of day-dependent gene expression is attenuated in arthritic joints, except for a subset of disease-modifying genes. Deletion of essential clock gene Bmal1 in FLS reduced susceptibility to collagen-induced arthritis (CIA), but did not impact symptomatic severity in affected mice. Notably, FLS Bmal1 deletion resulted in loss of diurnal expression of disease-modulating genes across the joint, and elevated production of MMP3, a prognostic marker of joint damage in inflammatory arthritis. This work identifies the FLS circadian clock as an influential driver of daily oscillations in joint inflammation, and a potential regulator of destructive pathology in chronic inflammatory arthritis.

immunology↗

Circadian clock function does not require the histone methyltransferase MLL3

AO_SCPLOWBSTRACTC_SCPLOWThe circadian clock controls the physiological function of tissues through the regulation of thousands of genes in a cell-type specific manner. The core cellular circadian clock is a transcription-translation negative feedback loop, which can recruit epigenetic regulators to facilitate temporal control of gene expression. Histone methyltransferase, mixed lineage leukemia gene 3 (MLL3) was reported to be required for maintenance of circadian oscillations in cultured cells. Here, we test the role of MLL3 in circadian organisation in whole animals. Using mice expressing catalytically inactive MLL3, we show that MLL3 methyltransferase activity is in fact not required for circadian oscillations in vitro in a range of tissues, nor for maintenance of circadian behavioural rhythms in vivo. In contrast to a previous report, loss of MLL3-dependent methylation did not affect global levels of H3K4 methylation in liver, indicating substantial compensation from other methyltransferases. Further, we found little evidence of genomic repositioning of H3K4me3 marks. We did, however, observe repositioning of H3K4me1 from intronic regions to intergenic regions and gene promoters, however there were no changes in H3K4me1 mark abundance around core circadian clock genes. Output functions of the circadian clock, such as control of inflammation, were largely intact in MLL3-methyltransferase deficient mice, although some gene specific changes were observed, with sexually dimorphic loss of circadian regulation of specific cytokines. Taken together, these observations call for a major reassessment of the inter-relationship between the circadian clock and MLL3-directed histone methylation, and a deeper examination of other epigenetic mechanisms which may facilitate circadian clock function. SO_SCPLOWIGNIFICANCEC_SCPLOWO_SCPCAP C_SCPCAPO_SCPLOWSTATEMENTC_SCPLOWA highly cited paper published in PNAS previously reported an essential role for the histone methyltransferase MLL3 in maintaining circadian oscillations in cultured cells. We tested the role of MLL3 in vivo and in primary tissues showing that MLL3 in fact plays no role in organising the core circadian clock, and has no functional impact on whole animal circadian behaviour. However, in further analysis, we newly discover a role for MLL3 in conferring circadian control to components of the inflammatory response, doing so in a sexually dimorphic manner. As the MLL family of histone methyltransferases are being targeted by pharmaceuticals for cancer, it is important to understand how methyltransferases may be driving circadian rhythms in gene expression.

molecular biology↗

HNF4A is required to specify glucocorticoid action in the liver

The glucocorticoid receptor (GR) is a nuclear hormone receptor critical to the regulation of energy metabolism and the inflammatory response. The actions of GR are highly dependent on cell type and environmental context. Here, we demonstrate the necessity for liver lineage-determining factor hepatocyte nuclear factor 4A (HNF4A) in defining liver-specificity of GR action. In normal mouse liver, the HNF4 motif lies adjacent to the glucocorticoid response element (GRE) at GR binding sites found within regions of open chromatin. In the absence of HNF4A, the liver GR cistrome is remodelled, with both loss and gain of GR recruitment evident. Lost sites are characterised by HNF4 motifs and weak GRE motifs. Gained sites are characterised by strong GRE motifs, and typically show GR recruitment in non-liver tissues. The functional importance of these HNF4A-regulated GR sites is further demonstrated by evidence of an altered transcriptional response to glucocorticoid treatment in the Hnf4a-null liver.

molecular biology↗

Adipocyte REVERBα dictates adipose tissue expansion during obesity

The circadian clock component REVERB is considered a dominant regulator of lipid metabolism, with global Reverb deletion driving dysregulation of white adipose tissue (WAT) lipogenesis and obesity. However, a similar phenotype is not observed under adipocyte-selective deletion (ReverbFlox2-6AdipoCre), and transcriptional profiling demonstrates that, under basal conditions, direct targets of REVERB regulation are limited, and include the circadian clock and collagen dynamics. Under high-fat diet (HFD) feeding, ReverbFlox2-6AdipoCre mice do manifest profound obesity, yet without the accompanying WAT inflammation and fibrosis exhibited by controls. Integration of the WAT REVERB cistrome with differential gene expression reveals broad control of metabolic processes by REVERB which is unmasked in the obese state. Adipocyte REVERB does not drive an anticipatory daily rhythm in WAT lipogenesis, but rather modulates WAT activity in response to alterations in metabolic state. Importantly, REVERB action in adipocytes is critical to the development of obesity-related WAT pathology and insulin resistance.

physiology↗

Ezh2 restrains macrophage inflammatory responses, and is critical for neutrophil migration in response to pulmonary infection.

Mucosal immunity is critical to survival, with huge attention at present due to the Coronovirus pandemic. Epigenetic factors are increasingly recognized as important determinants of immune responses, and EZH2 closest to application due to the availability of highly-specific and efficacious antagonists. However, very little is known about the role of EZH2 in the myeloid lineage, with some conflicting reports. Here we show EZH2 acts in macrophages to limit inflammatory responses to activation, and selective genetic deletion results in a remarkable gain in protection from infection with the prevalent lung pathogen, pneumococcus. In contrast, EZH2 is required for neutrophil chemotaxis, and animals lacking neutrophil EZH2 show increased susceptibility to pneumococcus. In summary, EZH2 shows complex, and divergent roles in different myeloid cells, likely contributing to the earlier conflicting reports. Compounds targeting EZH2 are likely to impair mucosal immunity, however, may prove useful for conditions driven by pulmonary neutrophil influx, such as adult respiratory distress syndrome (ARDS). DigestEpigenetic control of mucosal immunity is important, and has translational relevance with the advent of inhibitor drugs now in the clinic for cancer indications. Here we show divergent role for EZH2 in macrophages and neutrophils. Loss of EZH2 in macrophages results in a gain of inflammatory and immune function, and protection from pneumonia. However, EZH2 is required for neutrophil chemotaxis, resulting in impaired anti-bacterial defence. We show that inhibition, or loss of EZH2 in macrophages results in a gain of immune function, with increased responses to infectious mimics such as LPS. However, the impact was far more dramatic in-vivo, with striking protection from the consequences of infection with pneumococcal bacteria. Loss of EZH2 resulted in a gain in activity of a number of inflammatory signaling cascades, including NFkB, PPARg, and IRFs1, and 7. This widespread macrophage re-programming varied between macrophages sites of origin, with the greatest impact seen in peritoneal macrophages which resulted in emergence of a new population of MerTK low cells. In contrast, in the neutrophils loss of EZH2 greatly impairs motility, and chemotaxis. This results in dramatic impairment of immune responses to the same pneumococcal infection. Extension of these studies to the mucosal epithelium revealed that EZH2 in bronchoalveolar epithelial cells had no impact on responses to infection with influenza. Taken together EZH2 plays diverse roles in the myeloid lineage, with profound impacts on inflammatory responses. The most striking observation was the difference seen between macrophages and neutrophils. EZH2 inhibition is likely to greatly impair mucosal immunity. Impact StatementHere we show a striking, but highly cell-type specific impact of the EZH2 methyltransferase on inflammatory, and anti-infective circuits; inhibition of EZH2 in macrophages augments macrophage cytokine production, but by impairing neutrophil migration impairs anti-bacterial responses.

cell biology↗

The circadian clock protein REVERBα inhibits pulmonary fibrosis development

Pulmonary inflammatory responses lie under circadian control; however the importance of circadian mechanisms in fibrosis is not understood. Here, we identify a striking change to these mechanisms resulting in a gain of amplitude and lack of synchrony within pulmonary fibrotic tissue. These changes result from an infiltration of mesenchymal cells, an important cell type in the pathogenesis of pulmonary fibrosis. Mutation of the core clock protein REVERB in these cells exacerbated the development of bleomycin-induced fibrosis, whereas mutation of REVERB in club or myeloid cells had no effect on the bleomycin phenotype. Knockdown of REVERB revealed regulation of the poorly described transcription factor TBPL1. Both REVERB and TBPL1 altered integrin{beta}1 focal adhesion formation, resulting in increased myofibroblast activation. The translational importance of our findings was established through analysis of two human cohorts. In the UK Biobank circadian strain markers (sleep length, chronotype and shift work) are associated with pulmonary fibrosis making them novel risk factors. In a separate cohort REVERB expression was increased in human idiopathic pulmonary fibrosis (IPF) lung tissue. Pharmacological targeting of REVERB inhibited myofibroblast activation in IPF fibroblasts and collagen secretion in organotypic cultures from IPF patients, suggesting targeting REVERB could be a viable therapeutic approach.\n\nSignificanceThe circadian clock plays an essential role in energy metabolism, and inflammation. In contrast the importance of the clock in the pathogenesis of fibrosis remains poorly explored. This study describes a striking alteration in circadian biology during pulmonary fibrosis where the relatively arrhythmic alveolar structures gain circadian but desynchronous rhythmicity due to infiltration by fibroblasts. Disruption of the clock in these cells, which are not widely implicated in circadian pathophysiology, results in a pro-fibrotic phenotype. Translation of these findings in humans revealed previously unrecognised important circadian risk factors for pulmonary fibrosis (sleep length, chronotype and shift work). In addition, targeting REVERB repressed collagen secretion from human fibrotic lung tissue making this protein a promising therapeutic target.

cell biology↗

GWAS in 446,118 European adults identifies 78 genetic loci for self-reported habitual sleep duration supported by accelerometer-derived estimates

Sleep is an essential homeostatically-regulated state of decreased activity and alertness conserved across animal species, and both short and long sleep duration associate with chronic disease and all-cause mortality1,2. Defining genetic contributions to sleep duration could point to regulatory mechanisms and clarify causal disease relationships. Through genome-wide association analyses in 446,118 participants of European ancestry from the UK Biobank, we discover 78 loci for self-reported sleep duration that further impact accelerometer-derived measures of sleep duration, daytime inactivity duration, sleep efficiency and number of sleep bouts in a subgroup (n=85,499) with up to 7-day accelerometry. Associations are enriched for genes expressed in several brain regions, and for pathways including striatum and subpallium development, mechanosensory response, dopamine binding, synaptic neurotransmission, catecholamine production, synaptic plasticity, and unsaturated fatty acid metabolism. Genetic correlation analysis indicates shared biological links between sleep duration and psychiatric, cognitive, anthropometric and metabolic traits and Mendelian randomization highlights a causal link of longer sleep with schizophrenia.

genetics↗

Genome-wide association analyses of chronotype in 697,828 individuals provides new insights into circadian rhythms in humans and links to disease

Using genome-wide data from 697,828 research participants from 23andMe and UK Biobank, we increase the number of identified loci associated with being a morning person, a behavioural indicator of a persons underlying circadian rhythm, from 24 to 351. Using data from 85,760 individuals with activity-monitor derived measures of sleep timing we show that the chronotype loci influence sleep timing: the mean sleep timing of the 5% of individuals carrying the most "morningness" alleles was 25 minutes earlier than the 5% carrying the fewest. The loci were enriched for genes involved in circadian regulation, cAMP, glutamate and insulin signalling pathways, and those expressed in the retina, hindbrain, hypothalamus, and pituitary. We provide evidence that being a morning person is causally associated with better mental health but does not appear to affect BMI or Type 2 diabetes. This study offers new insights into the biology of circadian rhythms and links to disease in humans.

genetics↗

Biological and clinical insights from genetics of insomnia symptoms

Insomnia is a common disorder linked with adverse long-term medical and psychiatric outcomes, but underlying pathophysiological processes and causal relationships with disease are poorly understood. Here we identify 57 loci for self-reported insomnia symptoms in the UK Biobank (n=453,379) and confirm their impact on self-reported insomnia symptoms in the HUNT study (n=14,923 cases, 47,610 controls), physician diagnosed insomnia in Partners Biobank (n=2,217 cases, 14,240 controls), and accelerometer-derived measures of sleep efficiency and sleep duration in the UK Biobank (n=83,726). Our results suggest enrichment of genes involved in ubiquitin-mediated proteolysis, phototransduction and muscle development pathways and of genes expressed in multiple brain regions, skeletal muscle and adrenal gland. Evidence of shared genetic factors is found between frequent insomnia symptoms and restless legs syndrome, aging, cardio-metabolic, behavioral, psychiatric and reproductive traits. Evidence is found for a possible causal link between insomnia symptoms and coronary heart disease, depressive symptoms and subjective well-being.\n\nOne Sentence SummaryWe identify 57 genomic regions associated with insomnia pointing to the involvement of phototransduction and ubiquitination and potential causal links to CAD and depression.

genomics↗

Genome-wide association analyses of sleep disturbance traits identify new loci and highlight shared genetics with neuropsychiatric and metabolic traits

Chronic sleep disturbances, associated with cardio-metabolic diseases, psychiatric disorders and all-cause mortality1,2, affect 25-30% of adults worldwide3. While environmental factors contribute importantly to self-reported habitual sleep duration and disruption, these traits are heritable4-9, and gene identification should improve our understanding of sleep function, mechanisms linking sleep to disease, and development of novel therapies. We report single and multi-trait genome-wide association analyses (GWAS) of self-reported sleep duration, insomnia symptoms including difficulty initiating and/or maintaining sleep, and excessive daytime sleepiness in the UK Biobank (n=112,586), with discovery of loci for insomnia symptoms (near MEIS1, TMEM132E, CYCL1, TGFBI in females and WDR27 in males), excessive daytime sleepiness (near AR/OPHN1) and a composite sleep trait (near INADL and HCRTR2), as well as replication of a locus for sleep duration (at PAX-8). Genetic correlation was observed between longer sleep duration and schizophrenia (rG=0.29, p=1.90x10-13) and between increased excessive daytime sleepiness and increased adiposity traits (BMI rG=0.20, p=3.12x10-09; waist circumference rG=0.20, p=2.12x10-07).

genetics↗