Abstract

Smooth muscle cells (SMCs), the major cell type in atherosclerotic plaques, are vital in coronary artery diseases (CADs). Smooth muscle cell (SMC) phenotypic transition, which leads to the formation of various cell types in atherosclerotic plaques, is regulated by a network of genetic and epigenetic mechanisms and governs the risk of disease. The involvement of long noncoding RNAs (lncRNAs) has been increasingly identified in cardiovascular disease. However, SMC lncRNAs have not been comprehensively characterized, and their regulatory role in SMC state transition remains unknown.A discovery pipeline was constructed and applied to deeply strand-specific RNA sequencing from perturbed human coronary artery SMC with different disease-related stimuli, to allow for the detection of novel lncRNAs. The functional relevance of a select few novel lncRNAs were verified in vitro.We identified 4579 known and 13 655 de novo lncRNAs in human coronary artery SMC. Consistent with previous long noncoding RNA studies, these lncRNAs overall have fewer exons, are shorter in length than protein-coding genes (pcGenes), and have relatively low expression level. Genomic location of these long noncoding RNA is disproportionately enriched near CAD-related TFs (transcription factors), genetic loci, and gene regulators of SMC identity, suggesting the importance of their function in disease. Two de novo lncRNAs, ZEB-interacting suppressor (ZIPPOR) and TNS1-antisense (TNS1-AS2), were identified by our screen. Combining transcriptional data and in silico modeling along with in vitro validation, we identified CAD gene ZEB2 as a target through which these lncRNAs exert their function in SMC phenotypic transition.Expression of a large and diverse set of lncRNAs in human coronary artery SMC are highly dynamic in response to CAD-related stimuli. The dynamic changes in expression of these lncRNAs correspond to alterations in transcriptional programs that are relevant to CAD, suggesting a critical role for lncRNAs in SMC phenotypic transition and human atherosclerotic disease.

View details for DOI 10.1161/CIRCRESAHA.122.321960

View details for PubMedID 36852690

Abstract

Genome wide association studies for coronary artery disease (CAD) have identified a risk locus at 11q22.3. Here, we verify with mechanistic studies that rs2019090 and PDGFD represent the functional variant and gene at this locus. Further, FOXC1/C2 transcription factor binding at rs2019090 is shown to promote PDGFD transcription through the CAD promoting allele. With single cell transcriptomic and histology studies with Pdgfd knockdown in an SMC lineage tracing male atherosclerosis mouse model we find that Pdgfd promotes expansion, migration, and transition of SMC lineage cells to the chondromyocyte phenotype. Pdgfd also increases adventitial fibroblast and pericyte expression of chemokines and leukocyte adhesion molecules, which is linked to plaque macrophage recruitment. Despite these changes there is no effect of Pdgfd deletion on overall plaque burden. These findings suggest that PDGFD mediates CAD risk by promoting deleterious phenotypic changes in SMC, along with an inflammatory response that is primarily focused in the adventitia.

View details for DOI 10.1038/s41467-023-36518-9

View details for PubMedID 36792607

Abstract

To define the multi-cellular epigenomic and transcriptional landscape of cardiac cellular development, we generated single-cell chromatin accessibility maps of human fetal heart tissues. We identified eight major differentiation trajectories involving primary cardiac cell types, each associated with dynamic transcription factor (TF) activity signatures. We contrasted regulatory landscapes of iPSC-derived cardiac cell types and their invivo counterparts, which enabled optimization of invitro differentiation of epicardial cells. Further, we interpreted sequence based deep learning models of cell-type-resolved chromatin accessibility profiles to decipher underlying TF motif lexicons. De novo mutations predicted to affect chromatin accessibility in arterial endothelium were enriched in congenital heart disease (CHD) cases vs. controls. Invitro studies in iPSCs validated the functional impact of identified variation on the predicted developmental cell types. This work thus defines the cell-type-resolved cis-regulatory sequence determinants of heart development and identifies disruption of cell type-specific regulatory elements in CHD.

View details for DOI 10.1016/j.cell.2022.11.028

View details for PubMedID 36563664

Abstract

Atherosclerotic plaques consist mostly of smooth muscle cells (SMC), and genes that influence SMC phenotype can modulate coronary artery disease (CAD) risk. Allelic variation at 15q22.33 has been identified by genome-wide association studies to modify the risk of CAD and is associated with the expression of SMAD3 in SMC. However, the mechanism by which this gene modifies CAD risk remains poorly understood. Here we show that SMC-specific deletion of Smad3 in a murine atherosclerosis model resulted in greater plaque burden, more outward remodelling and increased vascular calcification. Single-cell transcriptomic analyses revealed that loss of Smad3 altered SMC transition cell state toward two fates: a SMC phenotype that governs both vascular remodelling and recruitment of inflammatory cells, as well as a chondromyocyte fate. Together, the findings reveal that Smad3 expression in SMC inhibits the emergence of specific SMC phenotypic transition cells that mediate adverse plaque features, including outward remodelling, monocyte recruitment, and vascular calcification.

View details for DOI 10.1038/s44161-022-00042-8

View details for PubMedID 36246779

View details for PubMedCentralID PMC9560061

Abstract

Background: Smooth muscle cells (SMC) transition into a number of different phenotypes during atherosclerosis, including those that resemble fibroblasts and chondrocytes, and make up the majority of cells in the atherosclerotic plaque. To better understand the epigenetic and transcriptional mechanisms that mediate these cell state changes, and how they relate to risk for coronary artery disease (CAD), we have investigated the causality and function of transcription factors (TFs) at genome wide associated loci. Methods: We employed CRISPR-Cas 9 genome and epigenome editing to identify the causal gene and cell(s) for a complex CAD GWAS signal at 2q22.3. Subsequently, single-cell epigenetic and transcriptomic profiling in murine models and human coronary artery smooth muscle cells were employed to understand the cellular and molecular mechanism by which this CAD risk gene exerts its function. Results: CRISPR-Cas 9 genome and epigenome editing showed that the complex CAD genetic signals within a genomic region at 2q22.3 lie within smooth muscle long-distance enhancers for ZEB2, a TF extensively studied in the context of epithelial mesenchymal transition (EMT) in development and cancer. ZEB2 regulates SMC phenotypic transition through chromatin remodeling that obviates accessibility and disrupts both Notch and TGFβ signaling, thus altering the epigenetic trajectory of SMC transitions. SMC specific loss of ZEB2 resulted in an inability of transitioning SMCs to turn off contractile programing and take on a fibroblast-like phenotype, but accelerated the formation of chondromyocytes, mirroring features of high-risk atherosclerotic plaques in human coronary arteries. Conclusions: These studies identify ZEB2 as a new CAD GWAS gene that affects features of plaque vulnerability through direct effects on the epigenome, providing a new thereapeutic approach to target vascular disease.

View details for DOI 10.1161/CIRCULATIONAHA.121.057789

Abstract

Background: Smooth muscle cells (SMC) transition into a number of different phenotypes during atherosclerosis, including those that resemble fibroblasts and chondrocytes, and make up the majority of cells in the atherosclerotic plaque. To better understand the epigenetic and transcriptional mechanisms that mediate these cell state changes, and how they relate to risk for coronary artery disease (CAD), we have investigated the causality and function of transcription factors (TFs) at genome wide associated loci. Methods: We employed CRISPR-Cas 9 genome and epigenome editing to identify the causal gene and cell(s) for a complex CAD GWAS signal at 2q22.3. Subsequently, single-cell epigenetic and transcriptomic profiling in murine models and human coronary artery smooth muscle cells were employed to understand the cellular and molecular mechanism by which this CAD risk gene exerts its function. Results: CRISPR-Cas 9 genome and epigenome editing showed that the complex CAD genetic signals within a genomic region at 2q22.3 lie within smooth muscle long-distance enhancers for ZEB2, a TF extensively studied in the context of epithelial mesenchymal transition (EMT) in development and cancer. ZEB2 regulates SMC phenotypic transition through chromatin remodeling that obviates accessibility and disrupts both Notch and TGFβ signaling, thus altering the epigenetic trajectory of SMC transitions. SMC specific loss of ZEB2 resulted in an inability of transitioning SMCs to turn off contractile programing and take on a fibroblast-like phenotype, but accelerated the formation of chondromyocytes, mirroring features of high-risk atherosclerotic plaques in human coronary arteries. Conclusions: These studies identify ZEB2 as a new CAD GWAS gene that affects features of plaque vulnerability through direct effects on the epigenome, providing a new thereapeutic approach to target vascular disease.

View details for DOI 10.1161/CIRCULATIONAHA.121.057789

View details for PubMedID 34990206

Abstract

To investigate the epigenetic and transcriptional mechanisms of coronary artery disease (CAD) risk, as well as the functional regulation of chromatin structure and function, we create a catalog of genetic variants associated with three stages of transcriptional cis-regulation in primary human coronary artery vascular smooth muscle cells (HCASMCs).We use a pooling approach with HCASMC lines to map regulatory variants that mediate binding of the CAD-associated transcription factor TCF21 with ChIPseq studies (bQTLs), variants that regulate chromatin accessibility with ATACseq studies (caQTLs), and chromosomal looping with Hi-C methods (clQTLs). We examine the overlap of these QTLs and their relationship to smooth muscle-specific genes and transcription factors. Further, we use multiple analyses to show that these QTLs are highly associated with CAD GWAS loci and correlate to lead SNPs where they show allelic effects. By utilizing genome editing, we verify that identified functional variants can regulate both chromatin accessibility and chromosomal looping, providing new insights into functional mechanisms regulating chromatin state and chromosomal structure. Finally, we directly link the disease-associated TGFB1-SMAD3 pathway to the CAD-associated FN1 gene through a response QTL that modulates both chromatin accessibility and chromosomal looping.Together, these studies represent the most thorough mapping of multiple QTL types in a highly disease-relevant primary cultured cell type and provide novel insights into their functional overlap and mechanisms that underlie these genomic features and their relationship to disease risk.

View details for DOI 10.1186/s13059-020-02049-5

View details for PubMedID 32513244

Abstract

Background: Smooth muscle cells (SMC) play a critical role in atherosclerosis. The Aryl hydrocarbon receptor (AHR) is an environment-sensing transcription factor that contributes to vascular development, and has been implicated in coronary artery disease (CAD) risk. We hypothesized that AHR can affect atherosclerosis by regulating phenotypic modulation of SMC. Methods: We combined RNA-Seq, ChIP-Seq, ATAC-Seq and in-vitro assays in human coronary artery SMC (HCASMC), with single-cell RNA-Seq (scRNA-Seq), histology, and RNAscope in an SMC-specific lineage-tracing Ahr knockout mouse model of atherosclerosis to better understand the role of AHR in vascular disease. Results: Genomic studies coupled with functional assays in cultured HCASMC revealed that AHR modulates HCASMC phenotype and suppresses ossification in these cells. Lineage tracing and activity tracing studies in the mouse aortic sinus showed that the Ahr pathway is active in modulated SMC in the atherosclerotic lesion cap. Furthermore, scRNA-Seq studies of the SMC-specific Ahr knockout mice showed a significant increase in the proportion of modulated SMC expressing chondrocyte markers such as Col2a1 and Alpl, which localized to the lesion neointima. These cells, which we term "chondromyocytes" (CMC), were also identified in the neointima of human coronary arteries. In histological analyses, these changes manifested as larger lesion size, increased lineage-traced SMC participation in the lesion, decreased lineage-traced SMC in the lesion cap, and increased alkaline phosphatase activity in lesions in the Ahr knockout compared to wild-type mice. We propose that AHR is likely protective based on these data and inference from human genetic analyses. Conclusions: Overall, we conclude that AHR promotes maintenance of lesion cap integrity and diminishes the disease related SMC-to-CMC transition in atherosclerotic tissues.

View details for DOI 10.1161/CIRCULATIONAHA.120.045981

View details for PubMedID 32441123

Abstract

Rationale: The gene encoding transcription factor TCF21 has been linked to coronary artery disease (CAD) risk by human genome wide association studies (GWAS) in multiple racial ethnic groups. In murine models, Tcf21 is required for phenotypic modulation of smooth muscle cells (SMC) in atherosclerotic tissues and promotes a fibroblast phenotype in these cells. In humans, TCF21 expression inhibits risk for CAD. The molecular mechanism by which TCF21 regulates SMC phenotype is not known. Objective: To better understand how TCF21 affects SMC phenotype, we sought to investigate the possible mechanisms by which it regulates the lineage determining myocardin (MYOCD)-serum response factor (SRF) pathway. Methods and Results: Modulation of TCF21 expression in HCASMC revealed that TCF21 suppresses a broad range of SMC markers, as well as key SMC transcription factors MYOCD and SRF, at the RNA and protein level. We conducted chromatin immunoprecipitation (ChIP)-sequencing to map SRF binding sites in HCASMC, showing that binding is colocalized in the genome with TCF21, including at a novel enhancer in the SRF gene, and at the MYOCD gene promoter. In vitro genome editing indicated that the SRF enhancer CArG box regulates transcription of the SRF gene, and mutation of this conserved motif in the orthologous mouse SRF enhancer revealed decreased SRF expression in aorta and heart tissues. Direct TCF21 binding and transcriptional inhibition at co-localized sites were established by reporter gene transfection assays. Chromatin immunoprecipitation and protein co-immunoprecipitation studies provided evidence that TCF21 blocks MYOCD and SRF association by direct TCF21-MYOCD interaction. Conclusions: These data indicate that TCF21 antagonizes the MYOCD-SRF pathway through multiple mechanisms, further establishing a role for this CAD associated gene in fundamental SMC processes and indicating the importance of smooth muscle response to vascular stress and phenotypic modulation of this cell type in CAD risk.

View details for DOI 10.1161/CIRCRESAHA.119.315968

View details for PubMedID 31815603

Abstract

In response to various stimuli, vascular smooth muscle cells (SMCs) can de-differentiate, proliferate and migrate in a process known as phenotypic modulation. However, the phenotype of modulated SMCs in vivo during atherosclerosis and the influence of this process on coronary artery disease (CAD) risk have not been clearly established. Using single-cell RNA sequencing, we comprehensively characterized the transcriptomic phenotype of modulated SMCs in vivo in atherosclerotic lesions of both mouse and human arteries and found that these cells transform into unique fibroblast-like cells, termed 'fibromyocytes', rather than into a classical macrophage phenotype. SMC-specific knockout of TCF21-a causal CAD gene-markedly inhibited SMC phenotypic modulation in mice, leading to the presence of fewer fibromyocytes within lesions as well as within the protective fibrous cap of the lesions. Moreover, TCF21 expression was strongly associated with SMC phenotypic modulation in diseased human coronary arteries, and higher levels of TCF21 expression were associated with decreased CAD risk in human CAD-relevant tissues. These results establish a protective role for both TCF21 and SMC phenotypic modulation in this disease.

View details for DOI 10.1038/s41591-019-0512-5

View details for PubMedID 31359001

View details for DOI 10.1186/s13073-019-0635-9

View details for Web of Science ID 000465465600001

Abstract

Coronary artery disease (CAD) is the leading cause of death globally. Genome-wide association studies (GWASs) have identified more than 95 independent loci that influence CAD risk, most of which reside in non-coding regions of the genome. To interpret these loci, we generated transcriptome and whole-genome datasets using human coronary artery smooth muscle cells (HCASMCs) from 52 unrelated donors, as well as epigenomic datasets using ATAC-seq on a subset of 8 donors. Through systematic comparison with publicly available datasets from GTEx and ENCODE projects, we identified transcriptomic, epigenetic, and genetic regulatory mechanisms specific to HCASMCs. We assessed the relevance of HCASMCs to CAD risk using transcriptomic and epigenomic level analyses. By jointly modeling eQTL and GWAS datasets, we identified five genes (SIPA1, TCF21, SMAD3, FES, and PDGFRA) that may modulate CAD risk through HCASMCs, all of which have relevant functional roles in vascular remodeling. Comparison with GTEx data suggests that SIPA1 and PDGFRA influence CAD risk predominantly through HCASMCs, while other annotated genes may have multiple cell and tissue targets. Together, these results provide tissue-specific and mechanistic insights into the regulation of a critical vascular cell type associated with CAD in human populations.

View details for PubMedID 30146127

View details for PubMedID 28818928

Abstract

The challenge of linking intergenic mutations to target genes has limited molecular understanding of human diseases. Here we show that H3K27ac HiChIP generates high-resolution contact maps of active enhancers and target genes in rare primary human T cell subtypes and coronary artery smooth muscle cells. Differentiation of naive T cells into T helper 17 cells or regulatory T cells creates subtype-specific enhancer-promoter interactions, specifically at regions of shared DNA accessibility. These data provide a principled means of assigning molecular functions to autoimmune and cardiovascular disease risk variants, linking hundreds of noncoding variants to putative gene targets. Target genes identified with HiChIP are further supported by CRISPR interference and activation at linked enhancers, by the presence of expression quantitative trait loci, and by allele-specific enhancer loops in patient-derived primary cells. The majority of disease-associated enhancers contact genes beyond the nearest gene in the linear genome, leading to a fourfold increase in the number of potential target genes for autoimmune and cardiovascular diseases.

View details for PubMedID 28945252

Abstract

Coronary artery disease (CAD) is the leading cause of mortality and morbidity, driven by both genetic and environmental risk factors. Meta-analyses of genome-wide association studies have identified >150 loci associated with CAD and myocardial infarction susceptibility in humans. A majority of these variants reside in non-coding regions and are co-inherited with hundreds of candidate regulatory variants, presenting a challenge to elucidate their functions. Herein, we use integrative genomic, epigenomic and transcriptomic profiling of perturbed human coronary artery smooth muscle cells and tissues to begin to identify causal regulatory variation and mechanisms responsible for CAD associations. Using these genome-wide maps, we prioritize 64 candidate variants and perform allele-specific binding and expression analyses at seven top candidate loci: 9p21.3, SMAD3, PDGFD, IL6R, BMP1, CCDC97/TGFB1 and LMOD1. We validate our findings in expression quantitative trait loci cohorts, which together reveal new links between CAD associations and regulatory function in the appropriate disease context.

View details for DOI 10.1038/ncomms12092

View details for PubMedID 27386823

View details for PubMedID 26837740

Abstract

TCF21 is a basic helix-loop-helix transcription factor that has recently been implicated as contributing to susceptibility to coronary heart disease based on genome wide association studies. In order to identify transcriptionally regulated target genes in a major disease relevant cell type, we performed siRNA knockdown of TCF21 in in vitro cultured human coronary artery smooth muscle cells and compared the transcriptome of siTCF21 versus siCONTROL treated cells. The raw (FASTQ) as well as processed (BED) data from 3 technical replicates per treatment has been deposited with Gene Expression Omnibus (GSE44461).

View details for PubMedID 26090325

Abstract

To functionally link coronary artery disease (CAD) causal genes identified by genome wide association studies (GWAS), and to investigate the cellular and molecular mechanisms of atherosclerosis, we have used chromatin immunoprecipitation sequencing (ChIP-Seq) with the CAD associated transcription factor TCF21 in human coronary artery smooth muscle cells (HCASMC). Analysis of identified TCF21 target genes for enrichment of molecular and cellular annotation terms identified processes relevant to CAD pathophysiology, including "growth factor binding," "matrix interaction," and "smooth muscle contraction." We characterized the canonical binding sequence for TCF21 as CAGCTG, identified AP-1 binding sites in TCF21 peaks, and by conducting ChIP-Seq for JUN and JUND in HCASMC confirmed that there is significant overlap between TCF21 and AP-1 binding loci in this cell type. Expression quantitative trait variation mapped to target genes of TCF21 was significantly enriched among variants with low P-values in the GWAS analyses, suggesting a possible functional interaction between TCF21 binding and causal variants in other CAD disease loci. Separate enrichment analyses found over-representation of TCF21 target genes among CAD associated genes, and linkage disequilibrium between TCF21 peak variation and that found in GWAS loci, consistent with the hypothesis that TCF21 may affect disease risk through interaction with other disease associated loci. Interestingly, enrichment for TCF21 target genes was also found among other genome wide association phenotypes, including height and inflammatory bowel disease, suggesting a functional profile important for basic cellular processes in non-vascular tissues. Thus, data and analyses presented here suggest that study of GWAS transcription factors may be a highly useful approach to identifying disease gene interactions and thus pathways that may be relevant to complex disease etiology.

View details for DOI 10.1371/journal.pgen.1005202

View details for Web of Science ID 000355305200022

View details for PubMedID 26020271

Abstract

Recent genome wide association studies have identified a number of genes that contribute to the risk for coronary heart disease. One such gene, TCF21, encodes a basic-helix-loop-helix transcription factor believed to serve a critical role in the development of epicardial progenitor cells that give rise to coronary artery smooth muscle cells (SMC) and cardiac fibroblasts. Using reporter gene and immunolocalization studies with mouse and human tissues we have found that vascular TCF21 expression in the adult is restricted primarily to adventitial cells associated with coronary arteries and also medial SMC in the proximal aorta of mouse. Genome wide RNA-Seq studies in human coronary artery SMC (HCASMC) with siRNA knockdown found a number of putative TCF21 downstream pathways identified by enrichment of terms related to CAD, including "vascular disease," "disorder of artery," and "occlusion of artery," as well as disease-related cellular functions including "cellular movement" and "cellular growth and proliferation." In vitro studies in HCASMC demonstrated that TCF21 expression promotes proliferation and migration and inhibits SMC lineage marker expression. Detailed in situ expression studies with reporter gene and lineage tracing revealed that vascular wall cells expressing Tcf21 before disease initiation migrate into vascular lesions of ApoE-/- and Ldlr-/- mice. While Tcf21 lineage traced cells are distributed throughout the early lesions, in mature lesions they contribute to the formation of a subcapsular layer of cells, and others become associated with the fibrous cap. The lineage traced fibrous cap cells activate expression of SMC markers and growth factor receptor genes. Taken together, these data suggest that TCF21 may have a role regulating the differentiation state of SMC precursor cells that migrate into vascular lesions and contribute to the fibrous cap and more broadly, in view of the association of this gene with human CAD, provide evidence that these processes may be a mechanism for CAD risk attributable to the vascular wall.

View details for DOI 10.1371/journal.pgen.1005155

View details for Web of Science ID 000355305200011

View details for PubMedID 26020946

Abstract

Recent genome wide association studies have identified a number of genes that contribute to the risk for coronary heart disease. One such gene, TCF21, encodes a basic-helix-loop-helix transcription factor believed to serve a critical role in the development of epicardial progenitor cells that give rise to coronary artery smooth muscle cells (SMC) and cardiac fibroblasts. Using reporter gene and immunolocalization studies with mouse and human tissues we have found that vascular TCF21 expression in the adult is restricted primarily to adventitial cells associated with coronary arteries and also medial SMC in the proximal aorta of mouse. Genome wide RNA-Seq studies in human coronary artery SMC (HCASMC) with siRNA knockdown found a number of putative TCF21 downstream pathways identified by enrichment of terms related to CAD, including "vascular disease," "disorder of artery," and "occlusion of artery," as well as disease-related cellular functions including "cellular movement" and "cellular growth and proliferation." In vitro studies in HCASMC demonstrated that TCF21 expression promotes proliferation and migration and inhibits SMC lineage marker expression. Detailed in situ expression studies with reporter gene and lineage tracing revealed that vascular wall cells expressing Tcf21 before disease initiation migrate into vascular lesions of ApoE-/- and Ldlr-/- mice. While Tcf21 lineage traced cells are distributed throughout the early lesions, in mature lesions they contribute to the formation of a subcapsular layer of cells, and others become associated with the fibrous cap. The lineage traced fibrous cap cells activate expression of SMC markers and growth factor receptor genes. Taken together, these data suggest that TCF21 may have a role regulating the differentiation state of SMC precursor cells that migrate into vascular lesions and contribute to the fibrous cap and more broadly, in view of the association of this gene with human CAD, provide evidence that these processes may be a mechanism for CAD risk attributable to the vascular wall.

View details for DOI 10.1371/journal.pgen.1005155

View details for PubMedID 26020946

Abstract

The role of long noncoding RNA (lncRNA) in adult hearts is unknown; also unclear is how lncRNA modulates nucleosome remodelling. An estimated 70% of mouse genes undergo antisense transcription, including myosin heavy chain 7 (Myh7), which encodes molecular motor proteins for heart contraction. Here we identify a cluster of lncRNA transcripts from Myh7 loci and demonstrate a new lncRNA-chromatin mechanism for heart failure. In mice, these transcripts, which we named myosin heavy-chain-associated RNA transcripts (Myheart, or Mhrt), are cardiac-specific and abundant in adult hearts. Pathological stress activates the Brg1-Hdac-Parp chromatin repressor complex to inhibit Mhrt transcription in the heart. Such stress-induced Mhrt repression is essential for cardiomyopathy to develop: restoring Mhrt to the pre-stress level protects the heart from hypertrophy and failure. Mhrt antagonizes the function of Brg1, a chromatin-remodelling factor that is activated by stress to trigger aberrant gene expression and cardiac myopathy. Mhrt prevents Brg1 from recognizing its genomic DNA targets, thus inhibiting chromatin targeting and gene regulation by Brg1. It does so by binding to the helicase domain of Brg1, a domain that is crucial for tethering Brg1 to chromatinized DNA targets. Brg1 helicase has dual nucleic-acid-binding specificities: it is capable of binding lncRNA (Mhrt) and chromatinized--but not naked--DNA. This dual-binding feature of helicase enables a competitive inhibition mechanism by which Mhrt sequesters Brg1 from its genomic DNA targets to prevent chromatin remodelling. A Mhrt-Brg1 feedback circuit is thus crucial for heart function. Human MHRT also originates from MYH7 loci and is repressed in various types of myopathic hearts, suggesting a conserved lncRNA mechanism in human cardiomyopathy. Our studies identify a cardioprotective lncRNA, define a new targeting mechanism for ATP-dependent chromatin-remodelling factors, and establish a new paradigm for lncRNA-chromatin interaction.

View details for DOI 10.1038/nature13596

View details for PubMedID 25119045

Abstract

Whole-genome sequencing (WGS) is increasingly applied in clinical medicine and is expected to uncover clinically significant findings regardless of sequencing indication.To examine coverage and concordance of clinically relevant genetic variation provided by WGS technologies; to quantitate inherited disease risk and pharmacogenomic findings in WGS data and resources required for their discovery and interpretation; and to evaluate clinical action prompted by WGS findings.An exploratory study of 12 adult participants recruited at Stanford University Medical Center who underwent WGS between November 2011 and March 2012. A multidisciplinary team reviewed all potentially reportable genetic findings. Five physicians proposed initial clinical follow-up based on the genetic findings.Genome coverage and sequencing platform concordance in different categories of genetic disease risk, person-hours spent curating candidate disease-risk variants, interpretation agreement between trained curators and disease genetics databases, burden of inherited disease risk and pharmacogenomic findings, and burden and interrater agreement of proposed clinical follow-up.Depending on sequencing platform, 10% to 19% of inherited disease genes were not covered to accepted standards for single nucleotide variant discovery. Genotype concordance was high for previously described single nucleotide genetic variants (99%-100%) but low for small insertion/deletion variants (53%-59%). Curation of 90 to 127 genetic variants in each participant required a median of 54 minutes (range, 5-223 minutes) per genetic variant, resulted in moderate classification agreement between professionals (Gross κ, 0.52; 95% CI, 0.40-0.64), and reclassified 69% of genetic variants cataloged as disease causing in mutation databases to variants of uncertain or lesser significance. Two to 6 personal disease-risk findings were discovered in each participant, including 1 frameshift deletion in the BRCA1 gene implicated in hereditary breast and ovarian cancer. Physician review of sequencing findings prompted consideration of a median of 1 to 3 initial diagnostic tests and referrals per participant, with fair interrater agreement about the suitability of WGS findings for clinical follow-up (Fleiss κ, 0.24; P < 001).In this exploratory study of 12 volunteer adults, the use of WGS was associated with incomplete coverage of inherited disease genes, low reproducibility of detection of genetic variation with the highest potential clinical effects, and uncertainty about clinically reportable findings. In certain cases, WGS will identify clinically actionable genetic variants warranting early medical intervention. These issues should be considered when determining the role of WGS in clinical medicine.

View details for DOI 10.1001/jama.2014.1717

View details for PubMedID 24618965

Abstract

Apelin and its cognate G protein-coupled receptor APJ constitute a signaling pathway with a positive inotropic effect on cardiac function and a vasodepressor function in the systemic circulation. The apelin-APJ pathway appears to have opposing physiological roles to the renin-angiotensin system. Here we investigated whether the apelin-APJ pathway can directly antagonize vascular disease-related Ang II actions. In ApoE-KO mice, exogenous Ang II induced atherosclerosis and abdominal aortic aneurysm formation; we found that coinfusion of apelin abrogated these effects. Similarly, apelin treatment rescued Ang II-mediated increases in neointimal formation and vascular remodeling in a vein graft model. NO has previously been implicated in the vasodepressor function of apelin; we found that apelin treatment increased NO bioavailability in ApoE-KO mice. Furthermore, infusion of an NO synthase inhibitor blocked the apelin-mediated decrease in atherosclerosis and aneurysm formation. In rat primary aortic smooth muscle cells, apelin inhibited Ang II-mediated transcriptional regulation of multiple targets as measured by reporter assays. In addition, we demonstrated by coimmunoprecipitation and fluorescence resonance energy transfer analysis that the Ang II and apelin receptors interacted physically. Taken together, these findings indicate that apelin signaling can block Ang II actions in vascular disease by increasing NO production and inhibiting Ang II cellular signaling.

View details for DOI 10.1172/JCI34871

View details for PubMedID 18769630