Estupiñán-Moreno E, Ortiz-Fernández L, Li T, Hernández-Rodríguez J, Ciudad L, Andrés-León E, Terron-Camero LC, Prieto-González S, Espígol-Frigolé G, Cid MC, Márquez A, Ballestar E, Martín J
Methylome and transcriptome profiling of giant cell arteritis monocytes reveals novel pathways involved in disease pathogenesis and molecular response to glucocorticoids.
Ann Rheum Dis15 Jun 2022, . Epub 15 Jun 2022
Giant cell arteritis (GCA) is a complex systemic vasculitis mediated by the interplay between both genetic and epigenetic factors. Monocytes are crucial players of the inflammation occurring in GCA. Therefore, characterisation of the monocyte methylome and transcriptome in GCA would be helpful to better understand disease pathogenesis.
Common variable immunodeficiency (CVID), the most prevalent symptomatic primary immunodeficiency, displays impaired terminal B-cell differentiation and defective antibody responses. Incomplete genetic penetrance and ample phenotypic expressivity in CVID suggest the participation of additional pathogenic mechanisms. Monozygotic (MZ) twins discordant for CVID are uniquely valuable for studying the contribution of epigenetics to the disease. Here, we generate a single-cell epigenomics and transcriptomics census of naïve-to-memory B cell differentiation in a CVID-discordant MZ twin pair. Our analysis identifies DNA methylation, chromatin accessibility and transcriptional defects in memory B-cells mirroring defective cell-cell communication upon activation. These findings are validated in a cohort of CVID patients and healthy donors. Our findings provide a comprehensive multi-omics map of alterations in naïve-to-memory B-cell transition in CVID and indicate links between the epigenome and immune cell cross-talk. Our resource, publicly available at the Human Cell Atlas, gives insight into future diagnosis and treatments of CVID patients.
de la Calle-Fabregat C, Rodríguez-Ubreva J, Ciudad L, Ramírez J, Celis R, Azuaga AB, Cuervo A, Graell E, Pérez-García C, Díaz-Torné C, Salvador G, Gómez-Puerta JA, Haro I, Sanmartí R, Cañete JD, Ballestar E
The synovial and blood monocyte DNA methylomes mirror prognosis, evolution and treatment in early arthritis.
JCI Insight24 Mar 2022, . Epub 24 Mar 2022
Identifying predictive biomarkers at early stages of early inflammatory arthritis is crucial for starting appropriate therapies to avoid poor outcomes. Monocytes and macrophages, largely associated with arthritis, are contributors and sensors of inflammation through epigenetic modifications. In this study, we investigated associations between clinical features and DNA methylation in blood and synovial fluid (SF) monocytes in a prospective cohort of early inflammatory arthritis patients. Undifferentiated arthritis (UA) blood monocyte DNA methylation profiles exhibited significant alterations in comparison with those from healthy donors. We identified additional differences both in blood and SF monocytes after comparing UA patients grouped by their future outcomes, good versus poor. Patient profiles in subsequent visits revealed a reversion towards a healthy level in both groups, those requiring disease-modifying antirheumatic drugs (DMARDs) and those that remitted spontaneously. Changes in disease activity between visits also impacted DNA methylation, partially concomitant in the SF of UA and in blood monocytes of rheumatoid arthritis patients. Epigenetic similarities between arthritis types allow a common prediction of disease activity. Our results constitute a resource of DNA methylation-based biomarkers of poor prognosis, disease activity and treatment efficacy in early untreated UA patients for the personalized clinical management of early inflammatory arthritis patients.
Català-Moll F, Ferreté-Bonastre AG, Godoy-Tena G, Morante-Palacios O, Ciudad L, Barberà L, Fondelli F, Martínez-Cáceres EM, Rodríguez-Ubreva J, Li T, Ballestar E
Vitamin D receptor, STAT3, and TET2 cooperate to establish tolerogenesis.
Cell Rep18 Jan 2022, 38(3)110244.
The active form of vitamin D, 1,25-dihydroxyvitamin D3, induces a stable tolerogenic phenotype in dendritic cells (DCs). This process involves the vitamin D receptor (VDR), which translocates to the nucleus, binds its cognate genomic sites, and promotes epigenetic and transcriptional remodeling. In this study, we report the occurrence of vitamin D-specific DNA demethylation and transcriptional activation at VDR binding sites associated with the acquisition of tolerogenesis in vitro. Differentiation to tolerogenic DCs associates with activation of the IL-6-JAK-STAT3 pathway. We show that JAK2-mediated STAT3 phosphorylation is specific to vitamin D stimulation. VDR and the phosphorylated form of STAT3 interact with each other to form a complex with methylcytosine dioxygenase TET2. Most importantly, pharmacological inhibition of JAK2 reverts vitamin D-induced tolerogenic properties of DCs. This interplay among VDR, STAT3, and TET2 opens up possibilities for modulating DC immunogenic properties in clinics.
Morante-Palacios O, Lorente-Sorolla C, Ciudad L, Calafell-Segura J, Garcia-Gomez A, Català-Moll F, Ruiz-Sanmartín A, Martínez-Gallo M, Ferrer R, Ruiz-Rodriguez JC, Álvarez-Errico D, Ballestar E
JAK2-STAT Epigenetically Regulates Tolerized Genes in Monocytes in the First Encounter With Gram-Negative Bacterial Endotoxins in Sepsis
Front. Immunol. 12:734652. https://doi.org/10.3389/fimmu.2021.73465217 Nov 2021, . Epub 17 Nov 2021
Microbial challenges, such as widespread bacterial infection in sepsis, induce endotoxin tolerance, a state of hyporesponsiveness to subsequent infections. The participation of DNA methylation in this process is poorly known. In this study, we perform integrated analysis of DNA methylation and transcriptional changes following in vitro exposure to gram-negative bacterial lipopolysaccharide, together with analysis of ex vivo monocytes from septic patients. We identify TET2-mediated demethylation and transcriptional activation of inflammation-related genes that is specific to toll-like receptor stimulation.
Changes also involve phosphorylation of STAT1, STAT3 and STAT5, elements of the JAK2 pathway. JAK2 pathway inhibition impairs the activation of tolerized genes on the first encounter with lipopolysaccharide. We then confirm the implication of the JAK2-STAT pathway in the aberrant DNA methylome of patients with sepsis caused by gram-negative bacteria. Finally, JAK2 inhibition in monocytes partially recapitulates the expression changes produced in the immunosuppressive cellular state acquired by monocytes from gram-negative sepsis, as described by single cell-RNA-sequencing. Our study evidences both the crucial role the JAK2-STAT pathway in epigenetic regulation and initial response of the tolerized genes to gram-negative bacterial endotoxins and provides a pharmacological target to prevent exacerbated responses.
Antonio Garcia-Gomez, Tianlu Li, Carlos de la Calle-Fabregat, Javier Rodríguez-Ubreva, Laura Ciudad, Francesc Català-Moll, Gerard Godoy-Tena, Montserrat Martín-Sánchez, Laura San-Segundo, Sandra Muntión, Xabier Morales, Carlos Ortiz-de-Solórzano, Julen Oyarzabal, Edurne San José-Enériz, Manel Esteller, Xabier Agirre, Felipe Prosper, Mercedes Garayoa, Esteban Ballestar
Targeting aberrant DNA methylation in mesenchymal stromal cells as a treatment for myeloma bone disease
Nat Commun 12, 421 (2021)18 Jan 2021, .
Multiple myeloma (MM) progression and myeloma-associated bone disease (MBD) are highly dependent on bone marrow mesenchymal stromal cells (MSCs). MM-MSCs exhibit abnormal transcriptomes, suggesting the involvement of epigenetic mechanisms governing their tumor-promoting functions and prolonged osteoblast suppression. Here, we identify widespread DNA methylation alterations of bone marrow-isolated MSCs from distinct MM stages, particularly in Homeobox genes involved in osteogenic differentiation that associate with their aberrant expression. Moreover, these DNA methylation changes are recapitulated in vitro by exposing MSCs from healthy individuals to MM cells. Pharmacological targeting of DNMTs and G9a with dual inhibitor CM-272 reverts the expression of hypermethylated osteogenic regulators and promotes osteoblast differentiation of myeloma MSCs. Most importantly, CM-272 treatment prevents tumor-associated bone loss and reduces tumor burden in a murine myeloma model. Our results demonstrate that epigenetic aberrancies mediate the impairment of bone formation in MM, and its targeting by CM-272 is able to reverse MBD.
Massoni-Badosa R, Iacono G, Moutinho C, Kulis M, Palau N, Marchese D, Rodríguez-Ubreva J, Ballestar E, Rodriguez-Esteban G, Marsal S, Aymerich M, Colomer D, Campo E, Julià A, Martín-Subero JI, Heyn H
Sampling time-dependent artifacts in single-cell genomics studies.
Genome Biol.11 May 2020, 21(1)112. Epub 11 May 2020
Robust protocols and automation now enable large-scale single-cell RNA and ATAC sequencing experiments and their application on biobank and clinical cohorts. However, technical biases introduced during sample acquisition can hinder solid, reproducible results, and a systematic benchmarking is required before entering large-scale data production. Here, we report the existence and extent of gene expression and chromatin accessibility artifacts introduced during sampling and identify experimental and computational solutions for their prevention.
Li, T., Ortiz, L., Andrés-León, E., Ciudad, L., Javierre, B.M., López-Isac, E., Guillén-Del-Castillo. A., Simeón-Aznar, C.P., Ballestar E, Martín, J.* / *co-corresponding/co-senior
Epigenomics and Transcriptomics of Systemic Sclerosis CD4+ T cells reveal Long Range Dysregulation of Key Inflammatory Pathways mediated by disease-associated Susceptibility Loci
Genome Medicine 12, 81 , .
Background: Systemic sclerosis (SSc) is a genetically complex autoimmune disease mediated by the interplay between genetic and epigenetic factors in a multitude of immune cells, with CD4+ T lymphocytes as one of the principle drivers of pathogenesis.
Methods: DNA samples exacted from CD4+ T cells of 48 SSc patients and 16 healthy controls were hybridized on MethylationEPIC BeadChip array. In parallel, gene expression was interrogated by hybridizing total RNA on Clariom™ S array. Downstream bioinformatics analyses were performed to identify correlating differentially methylated CpG positions (DMPs) and differentially expressed genes (DEGs), which were then confirmed utilizing previously published promoter capture Hi-C (PCHi-C) data.
Results: We identified 9112 and 3929 DMPs and DEGs, respectively. These DMPs and DEGs are enriched in functional categories related to inflammation and T cell biology. Furthermore, correlation analysis identified 17,500 possible DMP-DEG interaction pairs within a window of 5 Mb, and utilizing PCHi-C data, we observed that 212 CD4+ T cell-specific pairs of DMP-DEG also formed part of three-dimensional promoter-enhancer networks, potentially involving CTCF. Finally, combining PCHi-C data with SSc GWAS data, we identified four important SSc-associated susceptibility loci, TNIP1 (rs3792783), GSDMB (rs9303277), IL12RB1 (rs2305743), and CSK (rs1378942), that could potentially interact with DMP-DEG pairs cg17239269-ANXA6, cg19458020-CCR7, cg10808810-JUND, and cg11062629-ULK3, respectively.
Conclusion: Our study unveils a potential link between genetic, epigenetic, and transcriptional deregulation in CD4+ T cells of SSc patients, providing a novel integrated view of molecular components driving SSc pathogenesis.
Keywords: CTCF; DNA methylation; Epigenetics; Genetic susceptibility variants; Hi-C; Long-distance regulation; Systemic sclerosis.
Clinical value of DNA methylation markers in autoimmune rheumatic diseases.
Nature Reviews Rheumatology 16,514-524 , .
Methylation of cytosine residues in DNA, the best studied epigenetic modification, is associated with gene transcription and nuclear organization, and ultimately the function of a cell. DNA methylation can be influenced by various factors, including changes in neighbouring genomic sites such as those induced by transcription factor binding. The DNA methylation profiles in relevant cell types are altered in most human diseases compared with the healthy state. Given the physical stability of DNA and methylated DNA compared with other epigenetic modifications, DNA methylation is an ideal marker for clinical purposes. However, few DNA methylation-based markers have made it into clinical practice, with the notable exception of some markers used in the field of oncology. Autoimmune rheumatic diseases are genetically complex entities that can vary widely in terms of prognosis, subtypes, progression and treatment responses. Increasing reports showing strong links between DNA methylation profiles and different clinical outcomes and other clinical aspects in autoimmune rheumatic diseases reinforce the usefulness of DNA methylation profiles as novel clinical markers. In this Review, we provide an updated discussion on DNA methylation alterations in autoimmune rheumatic diseases and the advantages and disadvantages of using these markers in clinical practice.
Li, T., Garcia-Gomez, A., Morante-Palacios, O., Ciudad, L., Özkaramehmet, S., Van Dijck, E., Rodríguez-Ubreva, J., Vaquero, A. and, Ballestar E
SIRT1/2 orchestrate acquisition of DNA methylation and loss of histone H3 activating marks to prevent premature activation of inflammatory genes in macrophage
Nucleic Acids Research 48, 665–681 , .
Sirtuins 1 and 2 (SIRT1/2) are two NAD-dependent deacetylases with major roles in inflammation. In addition to deacetylating histones and other proteins, SIRT1/2-mediated regulation is coupled with other epigenetic enzymes. Here, we investigate the links between SIRT1/2 activity and DNA methylation in macrophage differentiation due to their relevance in myeloid cells. SIRT1/2 display drastic upregulation during macrophage differentiation and their inhibition impacts the expression of many inflammation-related genes. In this context, SIRT1/2 inhibition abrogates DNA methylation gains, but does not affect demethylation. Inhibition of hypermethylation occurs at many inflammatory loci, which results in more drastic upregulation of their expression upon macrophage polarization following bacterial lipopolysaccharide (LPS) challenge. SIRT1/2-mediated gains of methylation concur with decreases in activating histone marks, and their inhibition revert these histone marks to resemble an open chromatin. Remarkably, specific inhibition of DNA methyltransferases is sufficient to upregulate inflammatory genes that are maintained in a silent state by SIRT1/2. Both SIRT1 and SIRT2 directly interact with DNMT3B, and their binding to proinflammatory genes is lost upon exposure to LPS or through pharmacological inhibition of their activity. In all, we describe a novel role for SIRT1/2 to restrict premature activation of proinflammatory genes.