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• NCoR1 is the most highly expressed endothelial corepressor.
• Loss of NCoR1 promotes angiogenic function in endothelial cells.
• Loss of NCoR1 promotes a tip cell position during angiogenic sprouting.
Abstract
Corepressors negatively regulate gene expression by chromatin compaction. Targeted regulation of gene expression could provide a means to control endothelial cell phenotype. We hypothesize that by targeting corepressor proteins, endothelial angiogenic function can be improved. To study this, the expression and function of nuclear corepressors in human umbilical vein endothelial cells (HUVEC) and in murine organ culture was studied. RNA-seq revealed that nuclear receptor corepressor 1 (NCoR1), silencing mediator of retinoid and thyroid hormone receptors (SMRT) and repressor element-1 silencing transcription factor (REST) are the highest expressed corepressors in HUVECs. Knockout and knockdown strategies demonstrated that the depletion of NCoR1 increased the angiogenic capacity of endothelial cells, whereas depletion of SMRT or REST did not. Interestingly, the effect was VEGF signaling independent. NCoR1 depletion significantly upregulated angiogenesis-associated genes, especially tip cell genes, including ESM1, DLL4 and NOTCH4, as observed by RNA- and ATAC-seq. Confrontation assays comparing cells with and without NCoR1-deficiency revealed that loss of NCoR1 promotes a tip-cell position during spheroid sprouting. Moreover, a proximity ligation assay identified NCoR1 as a direct binding partner of the Notch-signaling-related transcription factor RBPJk. Luciferase assays showed that siRNA-mediated knockdown of NCOR1 promotes RBPJk activity. Furthermore, NCoR1 depletion prompts upregulation of several elements in the Notch signaling cascade. Downregulation of NOTCH4, but not NOTCH1, prevented the positive effect of NCOR1 knockdown on spheroid outgrowth. Collectively, these data indicate that decreasing NCOR1 expression is an attractive approach to promote angiogenic function.
Adhesion of human pathogenic bacteria to endothelial cells is facilitated by fibronectin interaction
(2023)
Human pathogenic bacteria circulating in the bloodstream need to find a way to interact with endothelial cells (ECs) lining the blood vessels to infect and colonise the host. The extracellular matrix (ECM) of ECs might represent an attractive initial target for bacterial interaction, as many bacterial adhesins have reported affinities to ECM proteins, in particular to fibronectin (Fn). Here, we analysed the general role of EC-expressed Fn for bacterial adhesion. For this, we evaluated the expression levels of ECM coding genes in different ECs, revealing that Fn is the highest expressed gene and thereby, it is highly abundant in the ECM environment of ECs. The role of Fn as a mediator in bacterial cell-host adhesion was evaluated in adhesion assays of Acinetobacter baumannii, Bartonella henselae, Borrelia burgdorferi, and Staphylococcus aureus to ECs. The assays demonstrated that bacteria colocalised with Fn fibres, as observed by confocal laser scanning microscopy. Fn removal from the ECM environment (FN1 knockout ECs) diminished bacterial adherence to ECs in both static and dynamic adhesion assays to varying extents, as evaluated via absolute quantification using qPCR. Interactions between adhesins and Fn might represent the crucial step for the adhesion of human-pathogenic Gram-negative and Gram-positive bacteria targeting the ECs as a niche of infection.
Macrophages are plastic and heterogeneous immune cells that adapt pro- or anti-inflammatory phenotypes upon exposure to different stimuli. Even though there has been evidence supporting a crosstalk between coagulation and innate immunity, the way in which protein components of the hemostasis pathway influence macrophages remains unclear. We investigated the effect of thrombin on macrophage polarization. On the basis of gene expression and cytokine secretion, our results suggest that polarization with thrombin induces an anti-inflammatory, M2-like phenotype. In functional studies, thrombin polarization promoted oxLDL phagocytosis by macrophages, and conditioned medium from the same cells increased endothelial cell proliferation. There were, however, clear differences between the classical M2a polarization and the effects of thrombin on gene expression. Finally, the deletion and inactivation of secreted modular Ca2+-binding protein 1 (SMOC1) attenuated phagocytosis by thrombin-stimulated macrophages, a phenomenon revered by the addition of recombinant SMOC1. Manipulation of SMOC1 levels also had a pronounced impact on the expression of TGF-β-signaling-related genes. Taken together, our results show that thrombin induces an anti-inflammatory macrophage phenotype with similarities as well as differences to the classical alternatively activated M2 polarization states, highlighting the importance of tissue levels of SMOC1 in modifying thrombin-induced macrophage polarization.
Long non-coding RNAs (lncRNAs) can act as regulatory RNAs which, by altering the expression of target genes, impact on the cellular phenotype and cardiovascular disease development. Endothelial lncRNAs and their vascular functions are largely undefined. Deep RNA-Seq and FANTOM5 CAGE analysis revealed the lncRNA LINC00607 to be highly enriched in human endothelial cells. LINC00607 was induced in response to hypoxia, arteriosclerosis regression in non-human primates and also in response to propranolol used to induce regression of human arteriovenous malformations. siRNA knockdown or CRISPR/Cas9 knockout of LINC00607 attenuated VEGF-A-induced angiogenic sprouting. LINC00607 knockout in endothelial cells also integrated less into newly formed vascular networks in an in vivo assay in SCID mice. Overexpression of LINC00607 in CRISPR knockout cells restored normal endothelial function. RNA- and ATAC-Seq after LINC00607 knockout revealed changes in the transcription of endothelial gene sets linked to the endothelial phenotype and in chromatin accessibility around ERG-binding sites. Mechanistically, LINC00607 interacted with the SWI/SNF chromatin remodeling protein BRG1. CRISPR/Cas9-mediated knockout of BRG1 in HUVEC followed by CUT&RUN revealed that BRG1 is required to secure a stable chromatin state, mainly on ERG-binding sites. In conclusion, LINC00607 is an endothelial-enriched lncRNA that maintains ERG target gene transcription by interacting with the chromatin remodeler BRG1.
Angiogenesis, the process by which endothelial cells (ECs) form new blood vessels from existing ones, is intimately linked to the tissue’s metabolic milieu and often occurs at nutrient-deficient sites. However, ECs rely on sufficient metabolic resources to support growth and proliferation. How endothelial nutrient acquisition and usage are regulated is unknown. Here we show that these processes are instructed by Yes-associated protein 1 (YAP)/WW domain-containing transcription regulator 1 (WWTR1/TAZ)-transcriptional enhanced associate domain (TEAD): a transcriptional module whose function is highly responsive to changes in the tissue environment. ECs lacking YAP/TAZ or their transcriptional partners, TEAD1, 2 and 4 fail to divide, resulting in stunted vascular growth in mice. Conversely, activation of TAZ, the more abundant paralogue in ECs, boosts proliferation, leading to vascular hyperplasia. We find that YAP/TAZ promote angiogenesis by fuelling nutrient-dependent mTORC1 signalling. By orchestrating the transcription of a repertoire of cell-surface transporters, including the large neutral amino acid transporter SLC7A5, YAP/TAZ-TEAD stimulate the import of amino acids and other essential nutrients, thereby enabling mTORC1 activation. Dissociating mTORC1 from these nutrient inputs—elicited by the loss of Rag GTPases—inhibits mTORC1 activity and prevents YAP/TAZ-dependent vascular growth. Together, these findings define a pivotal role for YAP/TAZ-TEAD in controlling endothelial mTORC1 and illustrate the essentiality of coordinated nutrient fluxes in the vasculature.
The transcription factor vitamin D receptor (VDR) is the high affinity nuclear target of the biologically active form of vitamin D3 (1,25(OH)2D3). In order to identify pure genomic transcriptional effects of 1,25(OH)2D3, we used VDR cistrome, transcriptome and open chromatin data, obtained from the human monocytic cell line THP-1, for a novel hierarchical analysis applying three bioinformatics approaches. We predicted 75.6% of all early 1,25(OH)2D3-responding (2.5 or 4 h) and 57.4% of the late differentially expressed genes (24 h) to be primary VDR target genes. VDR knockout led to a complete loss of 1,25(OH)2D3–induced genome-wide gene regulation. Thus, there was no indication of any VDR-independent non-genomic actions of 1,25(OH)2D3 modulating its transcriptional response. Among the predicted primary VDR target genes, 47 were coding for transcription factors and thus may mediate secondary 1,25(OH)2D3 responses. CEBPA and ETS1 ChIP-seq data and RNA-seq following CEBPA knockdown were used to validate the predicted regulation of secondary vitamin D target genes by both transcription factors. In conclusion, a directional network containing 47 partly novel primary VDR target transcription factors describes secondary responses in a highly complex vitamin D signaling cascade. The central transcription factor VDR is indispensable for all transcriptome-wide effects of the nuclear hormone.
Zinc finger proteins (ZNF) are a large group of transcription factors with diverse functions. We recently discovered that endothelial cells harbour a specific mechanism to limit the action of ZNF354C, whose function in endothelial cells is unknown. Given that ZNF354C has so far only been studied in bone and tumour, its function was determined in endothelial cells. ZNF354C is expressed in vascular cells and localises to the nucleus and cytoplasm. Overexpression of ZNF354C in human endothelial cells results in a marked inhibition of endothelial sprouting. RNA-sequencing of human microvascular endothelial cells with and without overexpression of ZNF354C revealed that the protein is a potent transcriptional repressor. ZNF354C contains an active KRAB domain which mediates this suppression as shown by mutagenesis analysis. ZNF354C interacts with dsDNA, TRIM28 and histones, as observed by proximity ligation and immunoprecipitation. Moreover, chromatin immunoprecipitation revealed that the ZNF binds to specific endothelial-relevant target-gene promoters. ZNF354C suppresses these genes as shown by CRISPR/Cas knockout and RNAi. Inhibition of endothelial sprouting by ZNF354C is dependent on the amino acids DV and MLE of the KRAB domain. These results demonstrate that ZNF354C is a repressive transcription factor which acts through a KRAB domain to inhibit endothelial angiogenic sprouting.
Despite a high clinical need for the treatment of colorectal carcinoma (CRC) as the second leading cause of cancer-related deaths, targeted therapies are still limited. The multifunctional enzyme Transglutaminase 2 (TGM2), which harbors transamidation and GTPase activity, has been implicated in the development and progression of different types of human cancers. However, the mechanism and role of TGM2 in colorectal cancer are poorly understood. Here, we present TGM2 as a promising drug target.
In primary patient material of CRC patients, we detected an increased expression and enzymatic activity of TGM2 in colon cancer tissue in comparison to matched normal colon mucosa cells. The genetic ablation of TGM2 in CRC cell lines using shRNAs or CRISPR/Cas9 inhibited cell expansion and tumorsphere formation. In vivo, tumor initiation and growth were reduced upon genetic knockdown of TGM2 in xenotransplantations. TGM2 ablation led to the induction of Caspase-3-driven apoptosis in CRC cells. Functional rescue experiments with TGM2 variants revealed that the transamidation activity is critical for the pro-survival function of TGM2. Transcriptomic and protein–protein interaction analyses applying various methods including super-resolution and time-lapse microscopy showed that TGM2 directly binds to the tumor suppressor p53, leading to its inactivation and escape of apoptosis induction.
We demonstrate here that TGM2 is an essential survival factor in CRC, highlighting the therapeutic potential of TGM2 inhibitors in CRC patients with high TGM2 expression. The inactivation of p53 by TGM2 binding indicates a general anti-apoptotic function, which may be relevant in cancers beyond CRC.
Endocannabinoids are important lipid-signaling mediators. Both protective and deleterious effects of endocannabinoids in the cardiovascular system have been reported but the mechanistic basis for these contradicting observations is unclear. We set out to identify anti-inflammatory mechanisms of endocannabinoids in the murine aorta and in human vascular smooth muscle cells (hVSMC). In response to combined stimulation with cytokines, IL-1β and TNFα, the murine aorta released several endocannabinoids, with anandamide (AEA) levels being the most significantly increased. AEA pretreatment had profound effects on cytokine-induced gene expression in hVSMC and murine aorta. As revealed by RNA-Seq analysis, the induction of a subset of 21 inflammatory target genes, including the important cytokine CCL2 was blocked by AEA. This effect was not mediated through AEA-dependent interference of the AP-1 or NF-κB pathways but rather through an epigenetic mechanism. In the presence of AEA, ATAC-Seq analysis and chromatin-immunoprecipitations revealed that CCL2 induction was blocked due to increased levels of H3K27me3 and a decrease of H3K27ac leading to compacted chromatin structure in the CCL2 promoter. These effects were mediated by recruitment of HDAC4 and the nuclear corepressor NCoR1 to the CCL2 promoter. This study therefore establishes a novel anti-inflammatory mechanism for the endogenous endocannabinoid AEA in vascular smooth muscle cells. Furthermore, this work provides a link between endogenous endocannabinoid signaling and epigenetic regulation.
Transcription factors can serve as links between tumor microenvironment signaling and oncogenesis. Interferon regulatory factor 9 (IRF9) is recruited and expressed upon interferon stimulation and is dependent on cofactors that exert in tumor-suppressing or oncogenic functions via the JAK-STAT pathway. IRF9 is frequently overexpressed in human lung cancer and is associated with decreased patient survival; however, the underlying mechanisms remain to be elucidated. Here, we used stably transduced lung adenocarcinoma cell lines (A549 and A427) to overexpress or knockdown IRF9. Overexpression led to increased oncogenic behavior in vitro, including enhanced proliferation and migration, whereas knockdown reduced these effects. These findings were confirmed in vivo using lung tumor xenografts in nude mice, and effects on both tumor growth and tumor mass were observed. Using RNA sequencing, we identified versican (VCAN) as a novel downstream target of IRF9. Indeed, IRF9 and VCAN expression levels were found to be correlated. We showed for the first time that IRF9 binds at a newly identified response element in the promoter region of VCAN to regulate its transcription. Using an siRNA approach, VCAN was found to enable the oncogenic properties (proliferation and migration) of IRF9 transduced cells, perhaps with CDKN1A involvement. The targeted inhibition of IRF9 in lung cancer could therefore be used as a new treatment option without multimodal interference in microenvironment JAK-STAT signaling.
Tumor-associated macrophages (TAMs) influence lung tumor development by inducing immunosuppression. Transcriptome analysis of TAMs isolated from human lung tumor tissues revealed an up-regulation of the Wnt/β-catenin pathway. These findings were reproduced in a newly developed in vitro “trained” TAM model. Pharmacological and macrophage-specific genetic ablation of β-catenin reprogrammed M2-like TAMs to M1-like TAMs both in vitro and in various in vivo models, which was linked with the suppression of primary and metastatic lung tumor growth. An in-depth analysis of the underlying signaling events revealed that β-catenin–mediated transcriptional activation of FOS-like antigen 2 (FOSL2) and repression of the AT-rich interaction domain 5A (ARID5A) drive gene regulatory switch from M1-like TAMs to M2-like TAMs. Moreover, we found that high expressions of β-catenin and FOSL2 correlated with poor prognosis in patients with lung cancer. In conclusion, β-catenin drives a transcriptional switch in the lung tumor microenvironment, thereby promoting tumor progression and metastasis.
With increasing distribution of endovascular stroke therapies, transient middle cerebral artery occlusion (tMCAO) in mice now more than ever depicts a relevant patient population with recanalized M1 occlusion. In this case, the desired therapeutic effect of blood flow restauration is accompanied by breakdown of the blood-brain barrier (BBB) and secondary reperfusion injury. The aim of this study was to elucidate short and intermediate-term transcriptional patterns and the involved pathways covering the different cellular players at the neurovascular unit after transient large vessel occlusion. To achieve this, male C57Bl/6J mice were treated according to an intensive post-stroke care protocol after 60 min occlusion of the middle cerebral artery or sham surgery to allow a high survival rate. After 24 h or 7 days, RNA from microvessel fragments from the ipsilateral and the contralateral hemispheres was isolated and used for mRNA sequencing. Bioinformatic analyses allowed us to depict gene expression changes at two timepoints of neurovascular post-stroke injury and regeneration. We validated our dataset by quantitative real time PCR of BBB-associated targets with well-characterized post-stroke dynamics. Hence, this study provides a well-controlled transcriptome dataset of a translationally relevant mouse model 24 h and 7 days after stroke which might help to discover future therapeutic targets in cerebral ischemia/reperfusion injury.
Protein aggregates and cytoplasmic vacuolization are major hallmarks of multisystem proteinopathies (MSPs) that lead to muscle weakness. Here, we identify METTL21C as a skeletal muscle-specific lysine methyltransferase. Insertion of a β-galactosidase cassette into the Mettl21c mouse locus revealed that METTL21C is specifically expressed in MYH7-positive skeletal muscle fibers. Ablation of the Mettl21c gene reduced endurance capacity and led to age-dependent accumulation of autophagic vacuoles in skeletal muscle. Denervation-induced muscle atrophy highlighted further impairments of autophagy-related proteins, including LC3, p62, and cathepsins, in Mettl21c−/− muscles. In addition, we demonstrate that METTL21C interacts with the ATPase p97 (VCP), which is mutated in various human MSP conditions. We reveal that METTL21C trimethylates p97 on the Lys315 residue and found that loss of this modification reduced p97 hexamer formation and ATPase activity in vivo. We conclude that the methyltransferase METTL21C is an important modulator of protein degradation in skeletal muscle under both normal and enhanced protein breakdown conditions.
Individual adult ventricular cardiomyocytes are either mono- or multi-nucleated and undergo morphological changes during cardiac hypertrophy. However, corresponding transcriptional signatures, reflecting potentially different functions or the ability for cell-cycle entry, are not known. The aim of this study was to determine the transcriptional profile of mono- and multi-nucleated adult cardiomyocytes by single-cell RNA-sequencing (scRNA-seq) and to investigate heterogeneity among cardiomyocytes under baseline conditions and in pressure-induced cardiac hypertrophy. We developed an array-based approach for scRNA-seq of rod-shaped multi-nucleated cardiomyocytes from both healthy and hypertrophic hearts. Single-cell transcriptomes of mono- or multi-nucleated cardiomyocytes were highly similar, although a certain degree of variation was noted across both populations. Non-image-based quality control allowing inclusion of damaged cardiomyocytes generated artificial cell clusters demonstrating the need for strict exclusion criteria. In contrast, cardiomyocytes isolated from hypertrophic heart after transverse aortic constriction showed heterogeneous transcriptional signatures, characteristic for hypoxia-induced responses. Immunofluorescence analysis revealed an inverse correlation between HIF1α+ cells and CD31-stained vessels, suggesting that imbalanced vascular growth in the hypertrophied heart induces cellular heterogeneity. Our study demonstrates that individual mono- and multi-nucleated cardiomyocytes express nearly identical sets of genes. Homogeneity among cardiomyocytes was lost after induction of hypertrophy due to differential HIF1α-dependent responses most likely caused by none-homogenous vessel growth.
Impaired alveolar formation and maintenance are features of many pulmonary diseases that are associated with significant morbidity and mortality. In a forward genetic screen for modulators of mouse lung development, we identified the non-muscle myosin II heavy chain gene, Myh10. Myh10 mutant pups exhibit cyanosis and respiratory distress, and die shortly after birth from differentiation defects in alveolar epithelium and mesenchyme. From omics analyses and follow up studies, we find decreased Thrombospondin expression accompanied with increased matrix metalloproteinase activity in both mutant lungs and cultured mutant fibroblasts, as well as disrupted extracellular matrix (ECM) remodeling. Loss of Myh10 specifically in mesenchymal cells results in ECM deposition defects and alveolar simplification. Notably, MYH10 expression is downregulated in the lung of emphysema patients. Altogether, our findings reveal critical roles for Myh10 in alveologenesis at least in part via the regulation of ECM remodeling, which may contribute to the pathogenesis of emphysema.
Formation and segregation of cell lineages forming the heart have been studied extensively but the underlying gene regulatory networks and epigenetic changes driving cell fate transitions during early cardiogenesis are still only partially understood. Here, we comprehensively characterize mouse cardiac progenitor cells (CPCs) marked by Nkx2-5 and Isl1 expression from E7.5 to E9.5 using single-cell RNA sequencing and transposase-accessible chromatin profiling (ATAC-seq). By leveraging on cell-to-cell transcriptome and chromatin accessibility heterogeneity, we identify different previously unknown cardiac subpopulations. Reconstruction of developmental trajectories reveal that multipotent Isl1+ CPC pass through an attractor state before separating into different developmental branches, whereas extended expression of Nkx2-5 commits CPC to an unidirectional cardiomyocyte fate. Furthermore, we show that CPC fate transitions are associated with distinct open chromatin states critically depending on Isl1 and Nkx2-5. Our data provide a model of transcriptional and epigenetic regulations during cardiac progenitor cell fate decisions at single-cell resolution.
G-protein-coupled receptor (GPCR) expression is extensively studied in bulk cDNA, but heterogeneity and functional patterning of GPCR expression in individual vascular cells is poorly understood. Here, we perform a microfluidic-based single-cell GPCR expression analysis in primary smooth muscle cells (SMC) and endothelial cells (EC). GPCR expression is highly heterogeneous in all cell types, which is confirmed in reporter mice, on the protein level and in human cells. Inflammatory activation in murine models of sepsis or atherosclerosis results in characteristic changes in the GPCR repertoire, and we identify functionally relevant subgroups of cells that are characterized by specific GPCR patterns. We further show that dedifferentiating SMC upregulate GPCRs such as Gpr39, Gprc5b, Gprc5c or Gpr124, and that selective targeting of Gprc5b modulates their differentiation state. Taken together, single-cell profiling identifies receptors expressed on pathologically relevant subpopulations and provides a basis for the development of new therapeutic strategies in vascular diseases.
Tubulogenesis is essential for the formation and function of internal organs. One such organ is the trachea, which allows gas exchange between the external environment and the lungs. However, the cellular and molecular mechanisms underlying tracheal tube development remain poorly understood. Here, we show that the potassium channel KCNJ13 is a critical modulator of tracheal tubulogenesis. We identify Kcnj13 in an ethylnitrosourea forward genetic screen for regulators of mouse respiratory organ development. Kcnj13 mutants exhibit a shorter trachea as well as defective smooth muscle (SM) cell alignment and polarity. KCNJ13 is essential to maintain ion homeostasis in tracheal SM cells, which is required for actin polymerization. This process appears to be mediated, at least in part, through activation of the actin regulator AKT, as pharmacological increase of AKT phosphorylation ameliorates the Kcnj13-mutant trachea phenotypes. These results provide insight into the role of ion homeostasis in cytoskeletal organization during tubulogenesis.
The mouse bitter taste receptors Tas2r143, Tas2r135, and Tas2r126 are encoded by genes that cluster on chromosome 6 and have been suggested to be expressed under common regulatory elements. Previous studies indicated that the Tas2r143/Tas2r135/Tas2r126 cluster is expressed in the heart, but other organs had not been systematically analyzed. In order to investigate the expression of this bitter taste receptor gene cluster in non-gustatory tissues, we generated a BAC (bacterial artificial chromosome) based transgenic mouse line, expressing CreERT2 under the control of the Tas2r143 promoter. After crossing this line with a mouse line expressing EGFP after Cre-mediated recombination, we were able to validate the Tas2r143-CreERT2 transgenic mouse line and monitor the expression of Tas2r143. EGFP-positive cells, indicating expression of members of the cluster, were found in about 47% of taste buds, and could also be found in several other organs. A population of EGFP-positive cells was identified in thymic epithelial cells, in the lamina propria of the intestine and in vascular smooth muscle cells of cardiac blood vessels. EGFP-positive cells were also identified in the epithelium of organs readily exposed to pathogens including lower airways, the gastrointestinal tract, urethra, vagina, and cervix. With respect to the function of cells expressing this bitter taste receptor cluster, RNA-seq analysis in EGFP-positive cells isolated from the epithelium of trachea and stomach showed expression of genes related to innate immunity. These data further support the concept that bitter taste receptors serve functions outside the gustatory system.
Background: The angiogenic function of endothelial cells is regulated by numerous mechanisms, but the impact of long noncoding RNAs (lncRNAs) has hardly been studied. We set out to identify novel and functionally important endothelial lncRNAs.
Methods: Epigenetically controlled lncRNAs in human umbilical vein endothelial cells were searched by exon-array analysis after knockdown of the histone demethylase JARID1B. Molecular mechanisms were investigated by RNA pulldown and immunoprecipitation, mass spectrometry, microarray, several knockdown approaches, CRISPR-Cas9, assay for transposase-accessible chromatin sequencing, and chromatin immunoprecipitation in human umbilical vein endothelial cells. Patient samples from lung and tumors were studied for MANTIS expression.
Results: A search for epigenetically controlled endothelial lncRNAs yielded lncRNA n342419, here termed MANTIS, as the most strongly regulated lncRNA. Controlled by the histone demethylase JARID1B, MANTIS was downregulated in patients with idiopathic pulmonary arterial hypertension and in rats treated with monocrotaline, whereas it was upregulated in carotid arteries of Macaca fascicularis subjected to atherosclerosis regression diet, and in endothelial cells isolated from human glioblastoma patients. CRISPR/Cas9-mediated deletion or silencing of MANTIS with small interfering RNAs or GapmeRs inhibited angiogenic sprouting and alignment of endothelial cells in response to shear stress. Mechanistically, the nuclear-localized MANTIS lncRNA interacted with BRG1, the catalytic subunit of the switch/sucrose nonfermentable chromatin-remodeling complex. This interaction was required for nucleosome remodeling by keeping the ATPase function of BRG1 active. Thereby, the transcription of key endothelial genes such as SOX18, SMAD6, and COUP-TFII was regulated by ensuring efficient RNA polymerase II machinery binding.
Conclusion: MANTIS is a differentially regulated novel lncRNA facilitating endothelial angiogenic function.