Suchergebnisse

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. ; This study was funded by: Spanish Ministry of Science and Innovation (grant number PID2020-117212RB-I00/AEI/10.13038/501100011033) (E.B.), Instituto de Salud Carlos III (ISCIII), Ref. AC18/00057, associated with i-PAD project (ERARE European Union program) (E.B.), Wellcome Sanger core funding (grant no. WT206194) (R.V.-T.), the Chan Zuckerberg Initiative (grant 2020-216799) (R.V.-T. and E.B.), an EMBO short-term fellowship (J.R.U.), Fondo de Investigación Sanitaria Instituto de Salud Carlos III (FIS PI16/01605) (L.P.-M.), the Spanish Ministry of Science, Innovation and Universities (SAF2017-89109-P; AEI/FEDER, UE) (H.H.), Instituto de Salud Carlos III, Ministry of Health (PI16/00759) and European Regional Development Fund-European Social Fund—FEDER-FSE) (C.R-G.), Grupo DISA (OA18/017) (C.R.-G.), the UK Biotechnology and Biological Sciences Research Council (BBS/E/B/000C0426) (G.K.) ...

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. ; HH is a Miguel Servet (CP14/00229) researcher funded by the Spanish Institute of Health Carlos III (ISCIII). CM and MK are supported by AECC postdoctoral fellowships. This work has received funding from the Ministerio de Ciencia, Innovación y Universidades (SAF2017-89109-P; AEI/FEDER, UE). This study was further funded by the Spanish Ministry of Economy and Competitiveness (grant number: IPT-010000-2010- 36, cofunded by the European Regional Development Fund). Core funding is from the ISCIII and the Generalitat de Catalunya. We acknowledge support of the Spanish Ministry of Economy, Industry and Competitiveness (MEIC) to the EMBL partnership, the Centro de Excelencia Severo Ochoa, the CERCA Programme/Generalitat de Catalunya, the Spanish Ministry of Economy, Industry and Competitiveness (MEIC) through the Instituto de Salud Carlos III and the Generalitat de Catalunya through Departament de Salut and Departament d'Empresa i Coneixement. We also acknowledge the Co-financing by the Spanish Ministry of Economy, Industry and Competitiveness (MEIC) with funds from the European Regional Development Fund (ERDF) corresponding to the 2014–2020 Smart Growth Operating Program. We acknowledge the Generalitat de Catalunya Suport Grups de Recerca AGAUR 2017-SGR-736 (to JIMS) and 2017-SGR-1142 (to EC), and CIBERONC (CB16/12/00225 and CB16/12/00334). EC is an ICREA Academia Researcher. This project received support from the European Commission under the projects DocTIS (H2020, SEP-210574908). This publication is part of a project (BCLLATLAS) that has received funding from the European Research Council (ERC) under the European Union's Horizon 2020 research and innovation program (grant agreement No 810287)

Abstract Background Epstein-Barr virus (EBV) infection is a well characterized etiopathogenic factor for a variety of immune-related conditions, including lymphomas, lymphoproliferative disorders and autoimmune diseases. EBV-mediated transformation of resting B cells to proliferating lymphoblastoid cells occurs in early stages of infection and is an excellent model for investigating the mechanisms associated with acquisition of unlimited growth. Results We investigated the effects of experimental EBV infection of B cells on DNA methylation profiles by using high-throughput analysis. Remarkably, we observed hypomethylation of around 250 genes, but no hypermethylation. Hypomethylation did not occur at repetitive sequences, consistent with the absence of genomic instability in lymphoproliferative cells. Changes in methylation only occurred after cell divisions started, without the participation of the active demethylation machinery, and were concomitant with acquisition by B cells of the ability to proliferate. Gene Ontology analysis, expression profiling, and high-throughput analysis of the presence of transcription factor binding motifs and occupancy revealed that most genes undergoing hypomethylation are active and display the presence of NF-κB p65 and other B cell-specific transcription factors. Promoter hypomethylation was associated with upregulation of genes relevant for the phenotype of proliferating lymphoblasts. Interestingly, pharmacologically induced demethylation increased the efficiency of transformation of resting B cells to lymphoblastoid cells, consistent with productive cooperation between hypomethylation and lymphocyte proliferation. Conclusions Our data provide novel clues on the role of the B cell transcription program leading to DNA methylation changes, which we find to be key to the EBV-associated conversion of resting B cells to proliferating lymphoblasts. ; This work was supported by grants PI081346 (FIS) and SAF2011-29635 from the Spanish Ministry of Science and Innovation (MICINN) and grant 2009SGR184 from AGAUR (Catalan Government). AI was supported by fellowship from AGAUR, Government of Catalonia, Spain. MP is supported by Ramon y Cajal Programme. ; Peer Reviewed

María Berdasco is with the Spanish National Cancer Centre, University of Oviedo, and Catalan Institute of Oncology; Rubén Alcázar is with University of Barcelona; María Victoria García-Ortiz is with University of Córdoba; Esteban Ballestar is with the Spanish National Cancer Centre; Agustín F. Fernández is with the Spanish National Cancer Centre; Teresa Roldán-Arjona is with University of Córdoba; Antonio F. Tiburcio is with University of Barcelona; Teresa Altabella is with University of Barcelona; Nicolas Buisine is with CNRS; Hadi Quesneville is with INRA; Antoine Baudry is with INRA; Loïc Lepiniec is with INRA; Miguel Alaminos is with University of Granada; Roberto Rodríguez is with University of Oviedo; Alan Lloyd is with UT Austin; Vincent Colot is with CNRS; Judith Bender is with Johns Hopkins University; María Jesús Canal is with University of Oviedo; Manel Esteller is with the Spanish National Cancer Centre and Catalan Institute of Oncology; Mario F. Fraga is with the Spanish National Cancer Centre and Spanish National Centre for Biotechnology. ; Maintaining and acquiring the pluripotent cell state in plants is critical to tissue regeneration and vegetative multiplication. Histone-based epigenetic mechanisms are important for regulating this undifferentiated state. Here we report the use of genetic and pharmacological experimental approaches to show that Arabidopsis cell suspensions and calluses specifically repress some genes as a result of promoter DNA hypermethylation. We found that promoters of the MAPK12, GSTU10 and BXL1 genes become hypermethylated in callus cells and that hypermethylation also affects the TTG1, GSTF5, SUVH8, fimbrin and CCD7 genes in cell suspensions. Promoter hypermethylation in undifferentiated cells was associated with histone hypoacetylation and primarily occurred at CpG sites. Accordingly, we found that the process specifically depends on MET1 and DRM2 methyltransferases, as demonstrated with DNA methyltransferase mutants. Our results suggest that promoter DNA methylation may be another important epigenetic mechanism for the establishment and/or maintenance of the undifferentiated state in plant cells. ; This work was supported by the Health (FIS01-04) (PI061267), Education and Science (I+D+I MCYT08-03, FU2004-02073/BMC and Consolider MEC09-05) Departments of the Spanish Government, the European Grant TRANSFOG LSHC-CT-2004-503438, and the Spanish Association Against Cancer (AECC). M.B. is funded by the Association Against Cancer (AECC). ; Cellular and Molecular Biology

This work was supported by the Health (FIS01-04) (PI061267), Education and Science (I+D+I MCYT08-03, FU2004-02073/BMC and Consolider MEC09-05) Departments of the Spanish Government, the European Grant TRANSFOG LSHC-CT-2004-503438, and the Spanish Association Against Cancer (AECC). M.B. is funded by the Association Against Cancer (AECC).

Maintaining and acquiring the pluripotent cell state in plants is critical to tissue regeneration and vegetative multiplication. Histone-based epigenetic mechanisms are important for regulating this undifferentiated state. Here we report the use of genetic and pharmacological experimental approaches to show that Arabidopsis cell suspensions and calluses specifically repress some genes as a result of promoter DNA hypermethylation. We found that promoters of the MAPK12, GSTU10 and BXL1 genes become hypermethylated in callus cells and that hypermethylation also affects the TTG1, GSTF5, SUVH8, fimbrin and CCD7 genes in cell suspensions. Promoter hypermethylation in undifferentiated cells was associated with histone hypoacetylation and primarily occurred at CpG sites. Accordingly, we found that the process specifically depends on MET1 and DRM2 methyltransferases, as demonstrated with DNA methyltransferase mutants. Our results suggest that promoter DNA methylation may be another important epigenetic mechanism for the establishment and/or maintenance of the undifferentiated state in plant cells. ; This work was supported by the Health (FIS01-04) (PI061267), Education and Science (I+D+I MCYT08-03, FU2004-02073/BMC and Consolider MEC09-05) Departments of the Spanish Government, the European Grant TRANSFOG LSHC-CT-2004-503438, and the Spanish Association Against Cancer (AECC). M.B. is funded by the Association Against Cancer (AECC).