Genome-wide chromatin contacts of super-enhancer-associated lncRNA identify LINC01013 as a regulator of fibrosis in the aortic valve.

Calcific aortic valve disease (CAVD) is characterized by a fibrocalcific process. The regulatory mechanisms that drive the fibrotic response in the aortic valve (AV) are poorly understood. Long noncoding RNAs derived from super-enhancers (lncRNA-SE) control gene expression and cell fate. Herein, multidimensional profiling including chromatin immunoprecipitation and sequencing, transposase-accessible chromatin sequencing, genome-wide 3D chromatin contacts of enhancer-promoter identified LINC01013 as an overexpressed lncRNA-SE during CAVD. LINC01013 is within a loop anchor, which has contact with the promoter of CCN2 (CTGF) located at ~180 kb upstream. Investigation showed that LINC01013 acts as a decoy factor for the negative transcription elongation factor E (NELF-E), whereby it controls the expression of CCN2. LINC01013-CCN2 is part of a transforming growth factor beta 1 (TGFB1) network and exerts a control over fibrogenesis. These findings illustrate a novel mechanism whereby a dysregulated lncRNA-SE controls, through a looping process, the expression of CCN2 and fibrogenesis of the AV.

Enhancer promoter interactome and Mendelian randomization identify network of druggable vascular genes in coronary artery disease.

Coronary artery disease (CAD) is a multifactorial disorder, which is partly heritable. Herein, we implemented a mapping of CAD-associated candidate genes by using genome-wide enhancer-promoter conformation (H3K27ac-HiChIP) and expression quantitative trait loci (eQTL). Enhancer-promoter anchor loops from human coronary artery smooth muscle cells (HCASMC) explained 22% of the heritability for CAD. 3D enhancer-promoter genome mapping of CAD-genes in HCASMC was enriched in vascular eQTL genes. By using colocalization and Mendelian randomization analyses, we identified 58 causal candidate vascular genes including some druggable targets (MAP3K11, CAMK1D, PDGFD, IPO9 and CETP). A network analysis of causal candidate genes was enriched in TGF beta and MAPK pathways. The pharmacologic inhibition of causal candidate gene MAP3K11 in vascular SMC reduced the expression of athero-relevant genes and lowered cell migration, a cardinal process in CAD. Genes connected to enhancers are enriched in vascular eQTL and druggable genes causally associated with CAD.

Oxyphospholipids in Cardiovascular Calcification.

Mineralization of cardiovascular structures including blood vessels and heart valves is a common feature. We postulate that ectopic mineralization is a response-to-injury in which signals delivered to cells trigger a chain of events to restore and repair tissues. Maladaptive response to external or internal signals promote the expression of danger-associated molecular patterns, which, in turn, promote, when expressed chronically, a procalcifying gene program. Growing evidence suggest that danger-associated molecular patterns such as oxyphospholipids and small lipid mediators, generated by enzyme activity, are involved in the transition of vascular smooth muscle cells and valve interstitial cells to an osteoblast-like phenotype. Understanding the regulation and the molecular processes underpinning the mineralization of atherosclerotic plaques and cardiac valves are providing valuable mechanistic insights, which could lead to the development of novel therapies. Herein, we provide a focus account on the role oxyphospholipids and their mediators in the development of mineralization in plaques and calcific aortic valve disease.

Enhancer-mediated enrichment of interacting JMJD3-DDX21 to ENPP2 locus prevents R-loop formation and promotes transcription.

ENPP2, which encodes for the enzyme autotaxin (ATX), is overexpressed during chronic inflammatory diseases and various cancers. However, the molecular mechanism involved in the ENPP2 transcription remains elusive. Here, in HEK 293T cells, we demonstrated that lipopolysaccharide (LPS) increased the transcription process at ENPP2 locus through a NF-кB pathway and a reduction of H3K27me3 level, a histone repressive mark, by the demethylase UTX. Simultaneously, the H3K27me3 demethylase JMJD3/KDM6B was recruited to the transcription start site (TSS), within the gene body and controlled the expression of ENPP2 in a non-enzymatic manner. Mass spectrometry data revealed a novel interaction for JMJD3 with DDX21, a RNA helicase that unwinds R-loops created by nascent transcript and DNA template. Upon LPS treatment, JMJD3 is necessary for DDX21 recruitment at ENPP2 locus allowing the resolution of aberrant R-loops. CRISPR-Cas9-mediated deletion of a distant-acting enhancer decreased the expression of ENPP2 and lowered the recruitment of JMJD3-DDX21 complex at TSS and its progression through the gene body. Taken together, these findings revealed that enhancer-mediated enrichment of novel JMJD3-DDX21 interaction at ENPP2 locus is necessary for nascent transcript synthesis via the resolution of aberrant R-loops formation in response to inflammatory stimulus.

System Genetics Including Causal Inference Identify Immune Targets for Coronary Artery Disease and the Lifespan.

BACKGROUND: Randomized clinical trials indicate that the immune response plays a significant role in coronary artery disease (CAD), a disorder impacting the lifespan potential. However, the identification of targets critical to the immune response in atheroma is still hampered by a lack of solid inference. METHODS: Herein, we implemented a system genetics approach to identify causally associated immune targets implicated in atheroma. We leveraged genome-wide association studies to perform mapping and Mendelian randomization to assess causal associations between gene expression in blood cells with CAD and the lifespan. Expressed genes (eGenes) were prioritized in network and in single-cell expression derived from plaque immune cells. RESULTS: Among 840 CAD-associated blood eGenes, 37 were predicted causally associated with CAD and 6 were also associated with the parental lifespan in Mendelian randomization. In multivariable Mendelian randomization, the impact of eGenes on the lifespan potential was mediated by the CAD risk. Predicted causal eGenes were central in network. FLT1 and CCR5 were identified as targets of approved drugs, whereas 22 eGenes were deemed tractable for the development of small molecules and antibodies. Analyses of plaque immune single-cell expression identified predicted causal eGenes enriched in macrophages (GPX1, C4orf3) and involved in ligand-receptor interactions (CCR5). CONCLUSIONS: We identified 37 blood eGenes predicted causally associated with CAD. The predicted expression for 6 eGenes impacted the lifespan potential through the risk of CAD. Prioritization based on network, annotations, and single-cell expression identified targets deemed tractable for the development of drugs and for drug repurposing.

Enhancer-associated aortic valve stenosis risk locus 1p21.2 alters NFATC2 binding site and promotes fibrogenesis.

Genome-wide association studies for calcific aortic valve stenosis (CAVS) previously reported strong signal for noncoding variants at 1p21.2. Previous study using Mendelian randomization suggested that the locus controls the expression of PALMD encoding Palmdelphin (PALMD). However, the molecular regulation at the locus and the impact of PALMD on the biology of the aortic valve is presently unknown. 3D genetic mapping and CRISPR activation identified rs6702619 as being located in a distant-acting enhancer, which controls the expression of PALMD. DNA-binding assay showed that the risk variant modified the DNA shape, which prevented the recruitment of NFATC2 and lowered the expression of PALMD. In co-expression network analysis, a module encompassing PALMD was enriched in actin-based process. Mass spectrometry and functional assessment showed that PALMD is a regulator of actin polymerization. In turn, lower level of PALMD promoted the activation of myocardin-related transcription factor and fibrosis, a key pathobiological process underpinning CAVS.

Prioritization of candidate causal genes for asthma in susceptibility loci derived from UK Biobank.

To identify candidate causal genes of asthma, we performed a genome-wide association study (GWAS) in UK Biobank on a broad asthma definition (n = 56,167 asthma cases and 352,255 controls). We then carried out functional mapping through transcriptome-wide association studies (TWAS) and Mendelian randomization in lung (n = 1,038) and blood (n = 31,684) tissues. The GWAS reveals 72 asthma-associated loci from 116 independent significant variants (PGWAS < 5.0E-8). The most significant lung TWAS gene on 17q12-q21 is GSDMB (PTWAS = 1.42E-54). Other TWAS genes include TSLP on 5q22, RERE on 1p36, CLEC16A on 16p13, and IL4R on 16p12, which all replicated in GTEx lung (n = 515). We demonstrate that the largest fold enrichment of regulatory and functional annotations among asthma-associated variants is in the blood. We map 485 blood eQTL-regulated genes associated with asthma and 50 of them are causal by Mendelian randomization. Prioritization of druggable genes reveals known (IL4R, TSLP, IL6, TNFSF4) and potentially new therapeutic targets for asthma.

Single-cell expression and Mendelian randomization analyses identify blood genes associated with lifespan and chronic diseases.

The human lifespan is a heritable trait, which is intricately linked to the development of disorders. Here, we show that genetic associations for the parental lifespan are enriched in open chromatin of blood cells. By using blood expression quantitative trait loci (eQTL) derived from 31,684 samples, we identified for the lifespan 125 cis- and 559 trans-regulated expressed genes (eGenes) enriched in adaptive and innate responses. Analysis of blood single-cell expression data showed that eGenes were enriched in dendritic cells (DCs) and the modelling of cell ligand-receptor interactions predicted crosstalk between DCs and a cluster of monocytes with a signature of cytotoxicity. In two-sample Mendelian randomization (MR), we identified 16 blood cis-eGenes causally associated with the lifespan. In MR, the majority of cis-eGene-disorder association pairs had concordant effects with the lifespan. The present work underlined that the lifespan is linked with the immune response and identifies eGenes associated with the lifespan and disorders.

A Mendelian randomization study of IL6 signaling in cardiovascular diseases, immune-related disorders and longevity.

Growing evidence suggests that inflammation is a significant contributor to different cardiovascular diseases (CVDs). Mendelian randomization (MR) was performed to assess the causal inference between plasma soluble IL6 receptor (sIL6R), a negative regulator of IL6 signaling, and different cardiovascular and immune-related disorders. Cis-MR with multiple instrumental variables showed an inverse association of sIL6R with rheumatoid arthritis, atrial fibrillation, stroke, coronary artery disease, and abdominal aortic aneurysm. However, genetically-determined sIL6R level was positively associated with atopic dermatitis and asthma. Also, sIL6R level was associated with longevity, as evaluated by parental age at death, a heritable trait. Gene-based association analysis with S-PrediXcan by using tissues from GTExV7 showed that IL6R tissue expression-disease pair associations were consistent with the directional effect of IL6 signaling identified in MR. Genetically-determined reduced IL6 signaling lowers the risk of multiple CVDs and is associated with increased longevity, but at the expense of higher atopic risk.