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1.
Nat Cardiovasc Res ; 3(9): 1067-1082, 2024 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-39271815

RESUMEN

Atrial fibrillation (AF) is the most common sustained arrhythmia and carries an increased risk of stroke and heart failure. Here we investigated how the immune infiltrate of human epicardial adipose tissue (EAT), which directly overlies the myocardium, contributes to AF. Flow cytometry analysis revealed an enrichment of tissue-resident memory T (TRM) cells in patients with AF. Cellular indexing of transcriptomes and epitopes by sequencing (CITE-seq) and single-cell T cell receptor (TCR) sequencing identified two transcriptionally distinct CD8+ TRM cells that are modulated in AF. Spatial transcriptomic analysis of EAT and atrial tissue identified the border region between the tissues to be a region of intense inflammatory and fibrotic activity, and the addition of TRM populations to atrial cardiomyocytes demonstrated their ability to differentially alter calcium flux as well as activate inflammatory and apoptotic signaling pathways. This study identified EAT as a reservoir of TRM cells that can directly modulate vulnerability to cardiac arrhythmia.


Asunto(s)
Tejido Adiposo , Fibrilación Atrial , Células T de Memoria , Pericardio , Fibrilación Atrial/inmunología , Fibrilación Atrial/genética , Fibrilación Atrial/patología , Fibrilación Atrial/metabolismo , Humanos , Pericardio/metabolismo , Pericardio/patología , Pericardio/inmunología , Tejido Adiposo/metabolismo , Tejido Adiposo/inmunología , Tejido Adiposo/patología , Células T de Memoria/inmunología , Células T de Memoria/metabolismo , Masculino , Linfocitos T CD8-positivos/inmunología , Linfocitos T CD8-positivos/metabolismo , Transcriptoma , Miocitos Cardíacos/metabolismo , Miocitos Cardíacos/patología , Miocitos Cardíacos/inmunología , Femenino , Persona de Mediana Edad , Perfilación de la Expresión Génica , Anciano , Fenotipo , Señalización del Calcio , Apoptosis , Memoria Inmunológica , Transcripción Genética , Estudios de Casos y Controles , Atrios Cardíacos/patología , Atrios Cardíacos/inmunología , Atrios Cardíacos/metabolismo , Fibrosis/patología , Tejido Adiposo Epicárdico
2.
Br J Haematol ; 204(3): 945-958, 2024 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-38296260

RESUMEN

EVI1 expression is associated with poor prognosis in myeloid leukaemia, which can result from Chr.3q alterations that juxtapose enhancers to induce EVI1 expression via long-range chromatin interactions. More often, however, EVI1 expression occurs unrelated to 3q alterations, and it remained unclear if, in these cases, EVI1 expression is similarly caused by aberrant enhancer activation. Here, we report that, in EVI1+3q- myeloid leukaemia cells, the EVI1 promoter interacts via long-range chromatin interactions with promoters of distally located, active genes, rather than with enhancer elements. Unlike in 3q+ cells, EVI1 expression and long-range interactions appear to not depend on CTCF/cohesin, though EVI1+3q- cells utilise an EVI1 promoter-proximal site to enhance its expression that is also involved in CTCF-mediated looping in 3q+ cells. Long-range interactions in 3q- cells connect EVI1 to promoters of multiple genes, whose transcription correlates with EVI1 in EVI1+3q- cell lines, suggesting a shared mechanism of transcriptional regulation. In line with this, CRISPR interference-induced silencing of two of these sites minimally, but consistently reduced EVI1 expression. Together, we provide novel evidence of features associated with EVI1 expression in 3q- leukaemia and consolidate the view that EVI1 in 3q- leukaemia is largely promoter-driven, potentially involving long-distance promoter clustering.


Asunto(s)
Leucemia Mieloide , Factores de Transcripción , Humanos , Factores de Transcripción/genética , Proteínas de Unión al ADN/genética , Cromatina , Proteína del Locus del Complejo MDS1 y EV11/genética , Leucemia Mieloide/genética , Proto-Oncogenes
3.
bioRxiv ; 2023 Mar 15.
Artículo en Inglés | MEDLINE | ID: mdl-36993619

RESUMEN

In most cell types, nuclear ß-catenin functions as prominent oncogenic driver and pairs with TCF7-family factors for transcriptional activation of MYC. Surprisingly, B-lymphoid malignancies not only lacked expression and activating lesions of ß-catenin but critically depended on GSK3ß for effective ß-catenin degradation. Our interactome studies in B-lymphoid tumors revealed that ß-catenin formed repressive complexes with lymphoid-specific Ikaros factors at the expense of TCF7. Instead of MYC-activation, ß-catenin was essential to enable Ikaros-mediated recruitment of nucleosome remodeling and deacetylation (NuRD) complexes for transcriptional repression of MYC. To leverage this previously unrecognized vulnerability of B-cell-specific repressive ß-catenin-Ikaros-complexes in refractory B-cell malignancies, we examined GSK3ß small molecule inhibitors to subvert ß-catenin degradation. Clinically approved GSK3ß-inhibitors that achieved favorable safety prof les at micromolar concentrations in clinical trials for neurological disorders and solid tumors were effective at low nanomolar concentrations in B-cell malignancies, induced massive accumulation of ß-catenin, repression of MYC and acute cell death. Preclinical in vivo treatment experiments in patient-derived xenografts validated small molecule GSK3ß-inhibitors for targeted engagement of lymphoid-specific ß-catenin-Ikaros complexes as a novel strategy to overcome conventional mechanisms of drug-resistance in refractory malignancies. HIGHLIGHTS: Unlike other cell lineages, B-cells express nuclear ß-catenin protein at low baseline levels and depend on GSK3ß for its degradation.In B-cells, ß-catenin forms unique complexes with lymphoid-specific Ikaros factors and is required for Ikaros-mediated tumor suppression and assembly of repressive NuRD complexes. CRISPR-based knockin mutation of a single Ikaros-binding motif in a lymphoid MYC superenhancer region reversed ß-catenin-dependent Myc repression and induction of cell death. The discovery of GSK3ß-dependent degradation of ß-catenin as unique B-lymphoid vulnerability provides a rationale to repurpose clinically approved GSK3ß-inhibitors for the treatment of refractory B-cell malignancies. GRAPHICAL ABSTRACT: Abundant nuclear ß-cateninß-catenin pairs with TCF7 factors for transcriptional activation of MYCB-cells rely on efficient degradation of ß-catenin by GSK3ßB-cell-specific expression of Ikaros factors Unique vulnerability in B-cell tumors: GSK3ß-inhibitors induce nuclear accumulation of ß-catenin.ß-catenin pairs with B-cell-specific Ikaros factors for transcriptional repression of MYC.

4.
Biomolecules ; 11(6)2021 06 07.
Artículo en Inglés | MEDLINE | ID: mdl-34200313

RESUMEN

Notch signaling forms an evolutionarily conserved juxtacrine pathway crucial for cellular development. Initially identified in Drosophila wing morphogenesis, Notch signaling has since been demonstrated to play pivotal roles in governing mammalian cellular development in a large variety of cell types. Indeed, abolishing Notch constituents in mouse models result in embryonic lethality, demonstrating that Notch signaling is critical for development and differentiation. In this review, we focus on the crucial role of Notch signaling in governing embryogenesis and differentiation of multiple progenitor cell types. Using hematopoiesis as a diverse cellular model, we highlight the role of Notch in regulating the cell fate of common lymphoid progenitors. Additionally, the influence of Notch through microenvironment interplay with lymphoid cells and how dysregulation influences disease processes is explored. Furthermore, bi-directional and lateral Notch signaling between ligand expressing source cells and target cells are investigated, indicating potentially novel therapeutic options for treatment of Notch-mediated diseases. Finally, we discuss the role of cis-inhibition in regulating Notch signaling in mammalian development.


Asunto(s)
Linaje de la Célula/fisiología , Desarrollo Embrionario/fisiología , Linfopoyesis/fisiología , Receptores Notch/fisiología , Animales , Humanos , Linfocitos/fisiología , Transducción de Señal/fisiología
5.
Mol Immunol ; 128: 150-164, 2020 12.
Artículo en Inglés | MEDLINE | ID: mdl-33129017

RESUMEN

During mammalian lymphoid development, Notch signaling is necessary at multiple stages of T lymphopoiesis, including lineage commitment, and later stages of T cell effector differentiation. In contrast, outside of a defined role in the development of splenic marginal zone B cells, there is conflicting evidence regarding whether Notch signaling plays functional roles in other B cell sub-populations. Complement receptor 2 (CR2) modulates BCR-signaling and is tightly regulated throughout differentiation. During B lymphopoiesis, CR2 is detected on immature and mature B cells with high surface expression on marginal zone B cells. Here, we have explored the possibility that Notch regulates human CR2 transcriptional activity using in vitro models including a co-culture system, co-transfection gene reporters and chromatin accessibility assays. We provide evidence that Notch signaling regulates CR2 promoter activity in a mature B cell line, as well as the induction of endogenous CR2 mRNA in a non-expressing pre-B cell line. The dynamics of endogenous gene activation suggests additional unidentified factors are required to mediate surface CR2 expression on immature and mature B lineage cells.


Asunto(s)
Complemento C3d/genética , Células Precursoras de Linfocitos B/fisiología , Regiones Promotoras Genéticas/genética , Receptores de Complemento 3d/genética , Receptores Notch/genética , Transducción de Señal/genética , Transcripción Genética/genética , Linfocitos B/fisiología , Diferenciación Celular/genética , Línea Celular , Línea Celular Tumoral , Cromatina/genética , Técnicas de Cocultivo/métodos , Humanos , Células K562 , Activación de Linfocitos/genética , Linfopoyesis/genética
6.
Hum Mutat ; 40(10): 1841-1855, 2019 10.
Artículo en Inglés | MEDLINE | ID: mdl-31112317

RESUMEN

The activities of DNA-binding transcription factors, such as the multi-zinc-finger protein ZBTB18 (also known as RP58, or ZNF238), are essential to coordinate mammalian neurodevelopment, including the birth and radial migration of newborn neurons within the fetal brain. In humans, the majority of disease-associated missense mutations in ZBTB18 lie within the DNA-binding zinc-finger domain and are associated with brain developmental disorder, yet the molecular mechanisms explaining their role in disease remain unclear. To address this, we developed in silico models of ZBTB18, bound to DNA, and discovered that half of the missense variants map to residues (Asn461, Arg464, Glu486) predicted to be essential to sequence-specific DNA contact, whereas others map to residues (Leu434, Tyr447, Arg495) with limited contributions to DNA binding. We studied pathogenic variants to residues with close (N461S) and limited (R495G) DNA contact and found that each bound DNA promiscuously, displayed altered transcriptional regulatory activity in vitro, and influenced the radial migration of newborn neurons in vivo in different ways. Taken together, our results suggest that altered transcriptional regulation could represent an important pathological mechanism for ZBTB18 missense variants in brain developmental disease.


Asunto(s)
Corteza Cerebral/embriología , Corteza Cerebral/metabolismo , Mutación Missense , Neuronas/metabolismo , Proteínas Represoras/genética , Dedos de Zinc/genética , Animales , Proteínas de Unión al ADN/química , Proteínas de Unión al ADN/metabolismo , Regulación de la Expresión Génica , Células HEK293 , Células HeLa , Humanos , Ratones , Modelos Moleculares , Unión Proteica , Conformación Proteica , Dominios y Motivos de Interacción de Proteínas , Proteínas Represoras/química , Relación Estructura-Actividad
7.
Ann Rheum Dis ; 75(1): 242-52, 2016 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-25180293

RESUMEN

OBJECTIVES: Systemic lupus erythematosus (SLE; OMIM 152700) is characterised by the production of antibodies to nuclear antigens. We previously identified variants in complement receptor 2 (CR2/CD21) that were associated with decreased risk of SLE. This study aimed to identify the causal variant for this association. METHODS: Genotyped and imputed genetic variants spanning CR2 were assessed for association with SLE in 15 750 case-control subjects from four ancestral groups. Allele-specific functional effects of associated variants were determined using quantitative real-time PCR, quantitative flow cytometry, electrophoretic mobility shift assay (EMSA) and chromatin immunoprecipitation (ChIP)-PCR. RESULTS: The strongest association signal was detected at rs1876453 in intron 1 of CR2 (pmeta=4.2×10(-4), OR 0.85), specifically when subjects were stratified based on the presence of dsDNA autoantibodies (case-control pmeta=7.6×10(-7), OR 0.71; case-only pmeta=1.9×10(-4), OR 0.75). Although allele-specific effects on B cell CR2 mRNA or protein levels were not identified, levels of complement receptor 1 (CR1/CD35) mRNA and protein were significantly higher on B cells of subjects harbouring the minor allele (p=0.0248 and p=0.0006, respectively). The minor allele altered the formation of several DNA protein complexes by EMSA, including one containing CCCTC-binding factor (CTCF), an effect that was confirmed by ChIP-PCR. CONCLUSIONS: These data suggest that rs1876453 in CR2 has long-range effects on gene regulation that decrease susceptibility to lupus. Since the minor allele at rs1876453 is preferentially associated with reduced risk of the highly specific dsDNA autoantibodies that are present in preclinical, active and severe lupus, understanding its mechanisms will have important therapeutic implications.


Asunto(s)
Anticuerpos Antinucleares/sangre , Lupus Eritematoso Sistémico/genética , Receptores de Complemento 3d/genética , Adolescente , Adulto , Subgrupos de Linfocitos B/inmunología , Estudios de Casos y Controles , ADN/inmunología , Predisposición Genética a la Enfermedad , Variación Genética , Genotipo , Haplotipos , Humanos , Lupus Eritematoso Sistémico/inmunología , Persona de Mediana Edad , Fenotipo , Polimorfismo de Nucleótido Simple , Receptores de Complemento 3b/biosíntesis , Medición de Riesgo/métodos , Factores de Transcripción/metabolismo , Adulto Joven
8.
Cell Mol Immunol ; 13(1): 119-31, 2016 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-25640655

RESUMEN

Complement receptor 2 (CR2/CD21) is predominantly expressed on the surface of mature B cells where it forms part of a coreceptor complex that functions, in part, to modulate B-cell receptor signal strength. CR2/CD21 expression is tightly regulated throughout B-cell development such that CR2/CD21 cannot be detected on pre-B or terminally differentiated plasma cells. CR2/CD21 expression is upregulated at B-cell maturation and can be induced by IL-4 and CD40 signaling pathways. We have previously characterized elements in the proximal promoter and first intron of CR2/CD21 that are involved in regulating basal and tissue-specific expression. We now extend these analyses to the CR2/CD21 core promoter. We show that in mature B cells, CR2/CD21 transcription proceeds from a focused TSS regulated by a non-consensus TATA box, an initiator element and a downstream promoter element. Furthermore, occupancy of the general transcriptional machinery in pre-B versus mature B-cell lines correlate with CR2/CD21 expression level and indicate that promoter accessibility must switch from inactive to active during the transitional B-cell window.


Asunto(s)
Antígenos CD40/metabolismo , Interleucina-4/metabolismo , Células Precursoras de Linfocitos B/metabolismo , Regiones Promotoras Genéticas , Receptores de Complemento 3d/metabolismo , Sitio de Iniciación de la Transcripción , Secuencia de Bases , Antígenos CD40/genética , Antígenos CD40/inmunología , Diferenciación Celular , Línea Celular Tumoral , Exones , Regulación de la Expresión Génica , Humanos , Interleucina-4/genética , Interleucina-4/inmunología , Intrones , Células K562 , Datos de Secuencia Molecular , Células Precursoras de Linfocitos B/citología , Células Precursoras de Linfocitos B/inmunología , Receptores de Complemento 3d/genética , Receptores de Complemento 3d/inmunología , Transducción de Señal , Transcripción Genética
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