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1.
Cell ; 184(15): 3915-3935.e21, 2021 07 22.
Artigo em Inglês | MEDLINE | ID: mdl-34174187

RESUMO

Emerging evidence indicates a fundamental role for the epigenome in immunity. Here, we mapped the epigenomic and transcriptional landscape of immunity to influenza vaccination in humans at the single-cell level. Vaccination against seasonal influenza induced persistently diminished H3K27ac in monocytes and myeloid dendritic cells (mDCs), which was associated with impaired cytokine responses to Toll-like receptor stimulation. Single-cell ATAC-seq analysis revealed an epigenomically distinct subcluster of monocytes with reduced chromatin accessibility at AP-1-targeted loci after vaccination. Similar effects were observed in response to vaccination with the AS03-adjuvanted H5N1 pandemic influenza vaccine. However, this vaccine also stimulated persistently increased chromatin accessibility at interferon response factor (IRF) loci in monocytes and mDCs. This was associated with elevated expression of antiviral genes and heightened resistance to the unrelated Zika and Dengue viruses. These results demonstrate that vaccination stimulates persistent epigenomic remodeling of the innate immune system and reveal AS03's potential as an epigenetic adjuvant.


Assuntos
Epigenômica , Imunidade/genética , Vacinas contra Influenza/genética , Vacinas contra Influenza/imunologia , Análise de Célula Única , Transcrição Gênica , Vacinação , Adolescente , Adulto , Antibacterianos/farmacologia , Antígenos CD34/metabolismo , Antivirais/farmacologia , Reprogramação Celular , Cromatina/metabolismo , Citocinas/biossíntese , Combinação de Medicamentos , Feminino , Regulação da Expressão Gênica , Histonas/metabolismo , Humanos , Imunidade Inata/genética , Virus da Influenza A Subtipo H5N1/efeitos dos fármacos , Virus da Influenza A Subtipo H5N1/imunologia , Interferon Tipo I/metabolismo , Masculino , Células Mieloides/metabolismo , Polissorbatos/farmacologia , Esqualeno/farmacologia , Receptores Toll-Like/metabolismo , Fator de Transcrição AP-1/metabolismo , Transcriptoma/genética , Adulto Jovem , alfa-Tocoferol/farmacologia
2.
Nat Immunol ; 22(6): 711-722, 2021 06.
Artigo em Inglês | MEDLINE | ID: mdl-34017121

RESUMO

Chromatin undergoes extensive reprogramming during immune cell differentiation. Here we report the repression of controlled histone H3 amino terminus proteolytic cleavage (H3ΔN) during monocyte-to-macrophage development. This abundant histone mark in human peripheral blood monocytes is catalyzed by neutrophil serine proteases (NSPs) cathepsin G, neutrophil elastase and proteinase 3. NSPs are repressed as monocytes mature into macrophages. Integrative epigenomic analysis reveals widespread H3ΔN distribution across the genome in a monocytic cell line and primary monocytes, which becomes largely undetectable in fully differentiated macrophages. H3ΔN is enriched at permissive chromatin and actively transcribed genes. Simultaneous NSP depletion in monocytic cells results in H3ΔN loss and further increase in chromatin accessibility, which likely primes the chromatin for gene expression reprogramming. Importantly, H3ΔN is reduced in monocytes from patients with systemic juvenile idiopathic arthritis, an autoinflammatory disease with prominent macrophage involvement. Overall, we uncover an epigenetic mechanism that primes the chromatin to facilitate macrophage development.


Assuntos
Artrite Juvenil/imunologia , Diferenciação Celular/imunologia , Epigênese Genética/imunologia , Histonas/metabolismo , Leucócitos Mononucleares/metabolismo , Macrófagos/imunologia , Adolescente , Artrite Juvenil/sangue , Artrite Juvenil/genética , Sistemas CRISPR-Cas/genética , Catepsina G/genética , Catepsina G/metabolismo , Diferenciação Celular/genética , Núcleo Celular/metabolismo , Criança , Pré-Escolar , Cromatina/metabolismo , Ensaios Enzimáticos , Epigenômica , Feminino , Técnicas de Inativação de Genes , Humanos , Células Jurkat , Elastase de Leucócito/genética , Elastase de Leucócito/metabolismo , Leucócitos Mononucleares/imunologia , Macrófagos/metabolismo , Masculino , Mieloblastina/genética , Mieloblastina/metabolismo , Cultura Primária de Células , Proteólise , RNA-Seq , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Células THP-1 , Adulto Jovem
3.
Cell ; 173(6): 1385-1397.e14, 2018 05 31.
Artigo em Inglês | MEDLINE | ID: mdl-29706550

RESUMO

Post-translational modifications of histone proteins and exchanges of histone variants of chromatin are central to the regulation of nearly all DNA-templated biological processes. However, the degree and variability of chromatin modifications in specific human immune cells remain largely unknown. Here, we employ a highly multiplexed mass cytometry analysis to profile the global levels of a broad array of chromatin modifications in primary human immune cells at the single-cell level. Our data reveal markedly different cell-type- and hematopoietic-lineage-specific chromatin modification patterns. Differential analysis between younger and older adults shows that aging is associated with increased heterogeneity between individuals and elevated cell-to-cell variability in chromatin modifications. Analysis of a twin cohort unveils heritability of chromatin modifications and demonstrates that aging-related chromatin alterations are predominantly driven by non-heritable influences. Together, we present a powerful platform for chromatin and immunology research. Our discoveries highlight the profound impacts of aging on chromatin modifications.


Assuntos
Envelhecimento , Cromatina/química , Epigênese Genética , Adolescente , Adulto , Idoso , Linhagem da Célula , Separação Celular , Doenças em Gêmeos , Feminino , Citometria de Fluxo , Histonas/metabolismo , Humanos , Sistema Imunitário , Imunofenotipagem , Leucócitos Mononucleares/citologia , Masculino , Pessoa de Meia-Idade , Monócitos/citologia , Análise de Componente Principal , Processamento de Proteína Pós-Traducional , Sistema de Registros , Adulto Jovem
4.
Nat Immunol ; 12(1): 29-36, 2011 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-21131967

RESUMO

Signaling via the methylation of lysine residues in proteins has been linked to diverse biological and disease processes, yet the catalytic activity and substrate specificity of many human protein lysine methyltransferases (PKMTs) are unknown. We screened over 40 candidate PKMTs and identified SETD6 as a methyltransferase that monomethylated chromatin-associated transcription factor NF-κB subunit RelA at Lys310 (RelAK310me1). SETD6-mediated methylation rendered RelA inert and attenuated RelA-driven transcriptional programs, including inflammatory responses in primary immune cells. RelAK310me1 was recognized by the ankryin repeat of the histone methyltransferase GLP, which under basal conditions promoted a repressed chromatin state at RelA target genes through GLP-mediated methylation of histone H3 Lys9 (H3K9). NF-κB-activation-linked phosphorylation of RelA at Ser311 by protein kinase C-ζ (PKC-ζ) blocked the binding of GLP to RelAK310me1 and relieved repression of the target gene. Our findings establish a previously uncharacterized mechanism by which chromatin signaling regulates inflammation programs.


Assuntos
Artrite Reumatoide/imunologia , NF-kappa B/metabolismo , Proteínas Metiltransferases/metabolismo , Fator de Transcrição RelA/metabolismo , Artrite Reumatoide/genética , Artrite Reumatoide/metabolismo , Montagem e Desmontagem da Cromatina/genética , Metilação de DNA , Células HEK293 , Histona-Lisina N-Metiltransferase/metabolismo , Histonas/metabolismo , Humanos , Inflamação , Lisina/metabolismo , NF-kappa B/genética , NF-kappa B/imunologia , Ligação Proteica/genética , Proteínas Metiltransferases/genética , Proteínas Metiltransferases/imunologia , RNA Interferente Pequeno/genética , Transdução de Sinais/genética , Transdução de Sinais/imunologia , Fator de Transcrição RelA/genética , Fator de Transcrição RelA/imunologia
5.
Genes Dev ; 28(16): 1758-71, 2014 Aug 15.
Artigo em Inglês | MEDLINE | ID: mdl-25128496

RESUMO

The dynamic reversible methylation of lysine residues on histone proteins is central to chromatin biology. Key components are demethylase enzymes, which remove methyl moieties from lysine residues. KDM2A, a member of the Jumonji C domain-containing histone lysine demethylase family, specifically targets lower methylation states of H3K36. Here, structural studies reveal that H3K36 specificity for KDM2A is mediated by the U-shaped threading of the H3K36 peptide through a catalytic groove within KDM2A. The side chain of methylated K36 inserts into the catalytic pocket occupied by Ni(2+) and cofactor, where it is positioned and oriented for demethylation. Key residues contributing to K36me specificity on histone H3 are G33 and G34 (positioned within a narrow channel), P38 (a turn residue), and Y41 (inserts into its own pocket). Given that KDM2A was found to also bind the H3K36me3 peptide, we postulate that steric constraints could prevent α-ketoglutarate from undergoing an "off-line"-to-"in-line" transition necessary for the demethylation reaction. Furthermore, structure-guided substitutions of residues in the KDM2A catalytic pocket abrogate KDM2A-mediated functions important for suppression of cancer cell phenotypes. Together, our results deduce insights into the molecular basis underlying KDM2A regulation of the biologically important methylated H3K36 mark.


Assuntos
Histonas/metabolismo , Histona Desmetilases com o Domínio Jumonji/química , Histona Desmetilases com o Domínio Jumonji/metabolismo , Modelos Moleculares , Animais , Sítios de Ligação , Linhagem Celular , Instabilidade Genômica , Histona Desmetilases com o Domínio Jumonji/genética , Metilação , Camundongos , Mutação , Ligação Proteica , Estrutura Quaternária de Proteína
6.
Mol Cell ; 50(3): 444-56, 2013 May 09.
Artigo em Inglês | MEDLINE | ID: mdl-23583077

RESUMO

Lysine methylation of histone proteins regulates chromatin dynamics and plays important roles in diverse physiological and pathological processes. However, beyond histone proteins, the proteome-wide extent of lysine methylation remains largely unknown. We have engineered the naturally occurring MBT domain repeats of L3MBTL1 to serve as a universal affinity reagent for detecting, enriching, and identifying proteins carrying a mono- or dimethylated lysine. The domain is broadly specific for methylated lysine ("pan-specific") and can be applied to any biological system. We have used our approach to demonstrate that SIRT1 is a substrate of the methyltransferase G9a both in vitro and in cells, to perform proteome-wide detection and enrichment of methylated proteins, and to identify candidate in-cell substrates of G9a and the related methyltransferase GLP. Together, our results demonstrate a powerful new approach for global and quantitative analysis of methylated lysine, and they represent the first systems biology understanding of lysine methylation.


Assuntos
Lisina/genética , Lisina/metabolismo , Proteoma/genética , Proteoma/metabolismo , Animais , Linhagem Celular , Células HEK293 , Humanos , Insetos , Metilação , Estrutura Terciária de Proteína , Proteômica/métodos , Células Sf9 , Sirtuína 1/genética , Sirtuína 1/metabolismo
7.
Mol Cell ; 44(4): 609-20, 2011 Nov 18.
Artigo em Inglês | MEDLINE | ID: mdl-22099308

RESUMO

The histone lysine methyltransferase NSD2 (MMSET/WHSC1) is implicated in diverse diseases and commonly overexpressed in multiple myeloma due to a recurrent t(4;14) chromosomal translocation. However, the precise catalytic activity of NSD2 is obscure, preventing progress in understanding how this enzyme influences chromatin biology and myeloma pathogenesis. Here, we show that dimethylation of histone H3 at lysine 36 (H3K36me2) is the principal chromatin-regulatory activity of NSD2. Catalysis of H3K36me2 by NSD2 is sufficient for gene activation. In t(4;14)-positive myeloma cells, the normal genome-wide and gene-specific distribution of H3K36me2 is obliterated, creating a chromatin landscape that selects for a transcription profile favorable for myelomagenesis. Catalytically active NSD2 confers xenograft tumor formation upon t(4;14)-negative cells and promotes oncogenic transformation of primary cells in an H3K36me2-dependent manner. Together, our findings establish H3K36me2 as the primary product generated by NSD2 and demonstrate that genomic disorganization of this canonical chromatin mark by NSD2 initiates oncogenic programming.


Assuntos
Transformação Celular Neoplásica , Regulação da Expressão Gênica , Histona-Lisina N-Metiltransferase , Histonas/metabolismo , Lisina/metabolismo , Mieloma Múltiplo/genética , Proteínas Recombinantes/metabolismo , Proteínas Repressoras , Transdução de Sinais/genética , Animais , Transformação Celular Neoplásica/genética , Transformação Celular Neoplásica/metabolismo , Cromatina , Perfilação da Expressão Gênica , Estudo de Associação Genômica Ampla , Histona-Lisina N-Metiltransferase/genética , Histona-Lisina N-Metiltransferase/metabolismo , Histonas/genética , Humanos , Metilação , Camundongos , Camundongos SCID , Mieloma Múltiplo/enzimologia , Mieloma Múltiplo/patologia , Proteínas Recombinantes/genética , Proteínas Repressoras/genética , Proteínas Repressoras/metabolismo , Transcrição Gênica , Translocação Genética , Ensaios Antitumorais Modelo de Xenoenxerto
8.
Clin Immunol ; 196: 40-48, 2018 11.
Artigo em Inglês | MEDLINE | ID: mdl-29960011

RESUMO

Modifications of histone proteins are fundamental to the regulation of epigenetic phenotypes. Dysregulations of histone modifications have been linked to the pathogenesis of diverse human diseases. However, identifying differential histone modifications in patients with immune-mediated diseases has been challenging, in part due to the lack of a powerful analytic platform to study histone modifications in the complex human immune system. We recently developed a highly multiplexed platform, Epigenetic landscape profiling using cytometry by Time-Of-Flight (EpiTOF), to analyze the global levels of a broad array of histone modifications in single cells using mass cytometry. In this review, we summarize the development of EpiTOF and discuss its potential applications in biomedical research. We anticipate that this platform will provide new insights into the roles of epigenetic regulation in hematopoiesis, immune cell functions, and immune system aging, and reveal aberrant epigenetic patterns associated with immune-mediated diseases.


Assuntos
Cromatina/metabolismo , Epigênese Genética , Código das Histonas , Histonas/metabolismo , Análise de Célula Única/métodos , Citometria de Fluxo , Humanos , Espectrometria de Massas , Processamento de Proteína Pós-Traducional
9.
Nature ; 484(7392): 115-9, 2012 Mar 07.
Artigo em Inglês | MEDLINE | ID: mdl-22398447

RESUMO

The recognition of distinctly modified histones by specialized 'effector' proteins constitutes a key mechanism for transducing molecular events at chromatin to biological outcomes. Effector proteins influence DNA-templated processes, including transcription, DNA recombination and DNA repair; however, no effector functions have yet been identified within the mammalian machinery that regulate DNA replication. Here we show that ORC1--a component of ORC (origin of replication complex), which mediates pre-DNA replication licensing--contains a bromo adjacent homology (BAH) domain that specifically recognizes histone H4 dimethylated at lysine 20 (H4K20me2). Recognition of H4K20me2 is a property common to BAH domains present within diverse metazoan ORC1 proteins. Structural studies reveal that the specificity of the BAH domain for H4K20me2 is mediated by a dynamic aromatic dimethyl-lysine-binding cage and multiple intermolecular contacts involving the bound peptide. H4K20me2 is enriched at replication origins, and abrogating ORC1 recognition of H4K20me2 in cells impairs ORC1 occupancy at replication origins, ORC chromatin loading and cell-cycle progression. Mutation of the ORC1 BAH domain has been implicated in the aetiology of Meier-Gorlin syndrome (MGS), a form of primordial dwarfism, and ORC1 depletion in zebrafish results in an MGS-like phenotype. We find that wild-type human ORC1, but not ORC1-H4K20me2-binding mutants, rescues the growth retardation of orc1 morphants. Moreover, zebrafish depleted of H4K20me2 have diminished body size, mirroring the phenotype of orc1 morphants. Together, our results identify the BAH domain as a novel methyl-lysine-binding module, thereby establishing the first direct link between histone methylation and the metazoan DNA replication machinery, and defining a pivotal aetiological role for the canonical H4K20me2 mark, via ORC1, in primordial dwarfism.


Assuntos
Replicação do DNA , Transtornos do Crescimento/metabolismo , Histonas/química , Histonas/metabolismo , Lisina/metabolismo , Micrognatismo/metabolismo , Complexo de Reconhecimento de Origem/química , Complexo de Reconhecimento de Origem/metabolismo , Animais , Proteínas de Transporte/genética , Proteínas de Transporte/metabolismo , Ciclo Celular , Linhagem Celular , Cromatina/genética , Cromatina/metabolismo , Microtia Congênita , Cristalografia por Raios X , Replicação do DNA/genética , Modelos Animais de Doenças , Nanismo/genética , Nanismo/metabolismo , Orelha/anormalidades , Transtornos do Crescimento/genética , Histonas/genética , Humanos , Metilação , Micrognatismo/genética , Modelos Moleculares , Complexo de Reconhecimento de Origem/genética , Patela/anormalidades , Patela/metabolismo , Estrutura Terciária de Proteína , Origem de Replicação , Peixe-Zebra , Proteínas de Peixe-Zebra/genética , Proteínas de Peixe-Zebra/metabolismo
10.
Nature ; 460(7251): 66-72, 2009 Jul 02.
Artigo em Inglês | MEDLINE | ID: mdl-19571879

RESUMO

Stem cells are controlled, in part, by genetic pathways frequently dysregulated during human tumorigenesis. Either stimulation of Wnt/beta-catenin signalling or overexpression of telomerase is sufficient to activate quiescent epidermal stem cells in vivo, although the mechanisms by which telomerase exerts these effects are not understood. Here we show that telomerase directly modulates Wnt/beta-catenin signalling by serving as a cofactor in a beta-catenin transcriptional complex. The telomerase protein component TERT (telomerase reverse transcriptase) interacts with BRG1 (also called SMARCA4), a SWI/SNF-related chromatin remodelling protein, and activates Wnt-dependent reporters in cultured cells and in vivo. TERT serves an essential role in formation of the anterior-posterior axis in Xenopus laevis embryos, and this defect in Wnt signalling manifests as homeotic transformations in the vertebrae of Tert(-/-) mice. Chromatin immunoprecipitation of the endogenous TERT protein from mouse gastrointestinal tract shows that TERT physically occupies gene promoters of Wnt-dependent genes. These data reveal an unanticipated role for telomerase as a transcriptional modulator of the Wnt/beta-catenin signalling pathway.


Assuntos
Cromatina/genética , Transdução de Sinais , Telomerase/metabolismo , Proteínas Wnt/metabolismo , Animais , Linhagem Celular , Coristoma/genética , Coristoma/patologia , DNA Helicases/metabolismo , Genes Reporter/genética , Células HeLa , Humanos , Intestino Delgado/metabolismo , Camundongos , Proteínas Nucleares/metabolismo , Oócitos/citologia , Oócitos/crescimento & desenvolvimento , Plasmídeos/genética , Regiões Promotoras Genéticas/genética , Somitos/anormalidades , Somitos/embriologia , Fatores de Transcrição/metabolismo , Proteínas Wnt/genética , Proteína Wnt3 , Xenopus laevis/embriologia , beta Catenina/genética
11.
Nature ; 452(7186): 492-6, 2008 Mar 27.
Artigo em Inglês | MEDLINE | ID: mdl-18337721

RESUMO

The Sir2 deacetylase regulates chromatin silencing and lifespan in Saccharomyces cerevisiae. In mice, deficiency for the Sir2 family member SIRT6 leads to a shortened lifespan and a premature ageing-like phenotype. However, the molecular mechanisms of SIRT6 function are unclear. SIRT6 is a chromatin-associated protein, but no enzymatic activity of SIRT6 at chromatin has yet been detected, and the identity of physiological SIRT6 substrates is unknown. Here we show that the human SIRT6 protein is an NAD+-dependent, histone H3 lysine 9 (H3K9) deacetylase that modulates telomeric chromatin. SIRT6 associates specifically with telomeres, and SIRT6 depletion leads to telomere dysfunction with end-to-end chromosomal fusions and premature cellular senescence. Moreover, SIRT6-depleted cells exhibit abnormal telomere structures that resemble defects observed in Werner syndrome, a premature ageing disorder. At telomeric chromatin, SIRT6 deacetylates H3K9 and is required for the stable association of WRN, the factor that is mutated in Werner syndrome. We propose that SIRT6 contributes to the propagation of a specialized chromatin state at mammalian telomeres, which in turn is required for proper telomere metabolism and function. Our findings constitute the first identification of a physiological enzymatic activity of SIRT6, and link chromatin regulation by SIRT6 to telomere maintenance and a human premature ageing syndrome.


Assuntos
Cromatina/metabolismo , Histona Desacetilases/metabolismo , Sirtuínas/metabolismo , Telômero/metabolismo , Acetilação , Linhagem Celular , Senescência Celular/genética , Cromatina/genética , Replicação do DNA , Exodesoxirribonucleases/metabolismo , Fibroblastos , Histona Desacetilases/deficiência , Histona Desacetilases/genética , Histonas/química , Histonas/metabolismo , Humanos , Lisina/metabolismo , Fenótipo , Ligação Proteica , RecQ Helicases/metabolismo , Sirtuínas/deficiência , Sirtuínas/genética , Telomerase/genética , Telomerase/metabolismo , Telômero/genética , Síndrome de Werner/genética , Helicase da Síndrome de Werner
12.
Nature ; 450(7172): 1106-10, 2007 Dec 13.
Artigo em Inglês | MEDLINE | ID: mdl-18033247

RESUMO

Nuclear processes such as transcription, DNA replication and recombination are dynamically regulated by chromatin structure. Eukaryotic transcription is known to be regulated by chromatin-associated proteins containing conserved protein domains that specifically recognize distinct covalent post-translational modifications on histones. However, it has been unclear whether similar mechanisms are involved in mammalian DNA recombination. Here we show that RAG2--an essential component of the RAG1/2 V(D)J recombinase, which mediates antigen-receptor gene assembly--contains a plant homeodomain (PHD) finger that specifically recognizes histone H3 trimethylated at lysine 4 (H3K4me3). The high-resolution crystal structure of the mouse RAG2 PHD finger bound to H3K4me3 reveals the molecular basis of H3K4me3-recognition by RAG2. Mutations that abrogate RAG2's recognition of H3K4me3 severely impair V(D)J recombination in vivo. Reducing the level of H3K4me3 similarly leads to a decrease in V(D)J recombination in vivo. Notably, a conserved tryptophan residue (W453) that constitutes a key structural component of the K4me3-binding surface and is essential for RAG2's recognition of H3K4me3 is mutated in patients with immunodeficiency syndromes. Together, our results identify a new function for histone methylation in mammalian DNA recombination. Furthermore, our results provide the first evidence indicating that disrupting the read-out of histone modifications can cause an inherited human disease.


Assuntos
Proteínas de Ligação a DNA/química , Proteínas de Ligação a DNA/metabolismo , Rearranjo Gênico do Linfócito B , Histonas/metabolismo , Lisina/metabolismo , Recombinação Genética , VDJ Recombinases/metabolismo , Motivos de Aminoácidos , Animais , Sítios de Ligação , Histonas/química , Proteínas de Homeodomínio/química , Proteínas de Homeodomínio/metabolismo , Humanos , Síndromes de Imunodeficiência/genética , Lisina/química , Metilação , Camundongos , Modelos Moleculares , Ligação Proteica , Relação Estrutura-Atividade , Especificidade por Substrato , Triptofano/genética , Triptofano/metabolismo , VDJ Recombinases/química
13.
J Crohns Colitis ; 17(5): 804-815, 2023 May 03.
Artigo em Inglês | MEDLINE | ID: mdl-36571819

RESUMO

BACKGROUND AND AIMS: Current understanding of histone post-translational modifications [histone modifications] across immune cell types in patients with inflammatory bowel disease [IBD] during remission and flare is limited. The present study aimed to quantify histone modifications at a single-cell resolution in IBD patients during remission and flare and how they differ compared to healthy controls. METHODS: We performed a case-control study of 94 subjects [83 IBD patients and 11 healthy controls]. IBD patients had either ulcerative colitis [n = 38] or Crohn's disease [n = 45] in clinical remission or flare. We used epigenetic profiling by time-of-flight [EpiTOF] to investigate changes in histone modifications within peripheral blood mononuclear cells from IBD patients. RESULTS: We discovered substantial heterogeneity in histone modifications across multiple immune cell types in IBD patients. They had a higher proportion of less differentiated CD34+ haematopoietic progenitors, and a subset of CD56bright natural killer [NK] cells and γδ T cells characterized by distinct histone modifications associated with gene transcription. The subset of CD56bright NK cells had increases in several histone acetylations. An epigenetically defined subset of NK cells was associated with higher levels of C-reactive protein in peripheral blood. CD34+ monocytes from IBD patients had significantly decreased cleaved H3T22, suggesting they were epigenetically primed for macrophage differentiation. CONCLUSION: We describe the first systems-level quantification of histone modifications across immune cells from IBD patients at a single-cell resolution, revealing the increased epigenetic heterogeneity that is not possible with traditional ChIP-seq profiling. Our data open new directions in investigating the association between histone modifications and IBD pathology using other epigenomic tools.


Assuntos
Colite Ulcerativa , Doenças Inflamatórias Intestinais , Humanos , Histonas/metabolismo , Leucócitos Mononucleares/metabolismo , Estudos de Casos e Controles , Processamento de Proteína Pós-Traducional
14.
JCI Insight ; 8(16)2023 08 22.
Artigo em Inglês | MEDLINE | ID: mdl-37606045

RESUMO

Systemic lupus erythematosus (SLE) affects 1 in 537 Black women, which is >2-fold more than White women. Black patients develop the disease at a younger age, have more severe symptoms, and have a greater chance of early mortality. We used a multiomics approach to uncover ancestry-associated immune alterations in patients with SLE and healthy controls that may contribute biologically to disease disparities. Cell composition, signaling, epigenetics, and proteomics were evaluated by mass cytometry; droplet-based single-cell transcriptomics and proteomics; and bead-based multiplex soluble mediator levels in plasma. We observed altered whole blood frequencies and enhanced activity in CD8+ T cells, B cells, monocytes, and DCs in Black patients with more active disease. Epigenetic modifications in CD8+ T cells (H3K27ac) could distinguish disease activity level in Black patients and differentiate Black from White patient samples. TLR3/4/7/8/9-related gene expression was elevated in immune cells from Black patients with SLE, and TLR7/8/9 and IFN-α phospho-signaling and cytokine responses were heightened even in immune cells from healthy Black control patients compared with White individuals. TLR stimulation of healthy immune cells recapitulated the ancestry-associated SLE immunophenotypes. This multiomic resource defines ancestry-associated immune phenotypes that differ between Black and White patients with SLE, which may influence the course and severity of SLE and other diseases.


Assuntos
Linfócitos B , Lúpus Eritematoso Sistêmico , Feminino , Humanos , População Negra , Linfócitos T CD8-Positivos , Lúpus Eritematoso Sistêmico/genética , Fenótipo , População Branca
15.
Nature ; 436(7053): 1048-52, 2005 Aug 18.
Artigo em Inglês | MEDLINE | ID: mdl-16107853

RESUMO

TERT, the protein component of telomerase, serves to maintain telomere function through the de novo addition of telomere repeats to chromosome ends, and is reactivated in 90% of human cancers. In normal tissues, TERT is expressed in stem cells and in progenitor cells, but its role in these compartments is not fully understood. Here we show that conditional transgenic induction of TERT in mouse skin epithelium causes a rapid transition from telogen (the resting phase of the hair follicle cycle) to anagen (the active phase), thereby facilitating robust hair growth. TERT overexpression promotes this developmental transition by causing proliferation of quiescent, multipotent stem cells in the hair follicle bulge region. This new function for TERT does not require the telomerase RNA component, which encodes the template for telomere addition, and therefore operates through a mechanism independent of its activity in synthesizing telomere repeats. These data indicate that, in addition to its established role in extending telomeres, TERT can promote proliferation of resting stem cells through a non-canonical pathway.


Assuntos
Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Folículo Piloso/citologia , Células-Tronco/citologia , Telomerase/genética , Telomerase/metabolismo , Animais , Bromodesoxiuridina/metabolismo , Proliferação de Células , Proteínas de Ligação a DNA/deficiência , Indução Enzimática , Folículo Piloso/enzimologia , Camundongos , Camundongos Transgênicos , RNA/genética , Células-Tronco/enzimologia , Telomerase/deficiência , Transgenes/genética
16.
PLoS Genet ; 4(1): e10, 2008 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-18208333

RESUMO

Telomerase serves a critical role in stem cell function and tissue homeostasis. This role depends on its ability to synthesize telomere repeats in a manner dependent on the reverse transcriptase (RT) function of its protein component telomerase RT (TERT), as well as on a novel pathway whose mechanism is poorly understood. Here, we use a TERT mutant lacking RT function (TERT(ci)) to study the mechanism of TERT action in mammalian skin, an ideal tissue for studying progenitor cell biology. We show that TERT(ci) retains the full activities of wild-type TERT in enhancing keratinocyte proliferation in skin and in activating resting hair follicle stem cells, which triggers initiation of a new hair follicle growth phase and promotes hair synthesis. To understand the nature of this RT-independent function for TERT, we studied the genome-wide transcriptional response to acute changes in TERT levels in mouse skin. We find that TERT facilitates activation of progenitor cells in the skin and hair follicle by triggering a rapid change in gene expression that significantly overlaps the program controlling natural hair follicle cycling in wild-type mice. Statistical comparisons to other microarray gene sets using pattern-matching algorithms revealed that the TERT transcriptional response strongly resembles those mediated by Myc and Wnt, two proteins intimately associated with stem cell function and cancer. These data show that TERT controls tissue progenitor cells via transcriptional regulation of a developmental program converging on the Myc and Wnt pathways.


Assuntos
Células Epiteliais/fisiologia , Regulação da Expressão Gênica no Desenvolvimento , Proteínas Proto-Oncogênicas c-myc/genética , Telomerase/metabolismo , Proteínas Wnt/genética , Algoritmos , Animais , Biópsia , Proliferação de Células , Procedimentos Cirúrgicos Dermatológicos , Perfilação da Expressão Gênica , Cabelo/metabolismo , Folículo Piloso/citologia , Folículo Piloso/metabolismo , Imuno-Histoquímica , Queratinócitos/fisiologia , Camundongos , Camundongos Transgênicos , Modelos Biológicos , Mutação , RNA Mensageiro/metabolismo , Pele/citologia , Pele/enzimologia , Pele/metabolismo , Células-Tronco/citologia , Células-Tronco/metabolismo , Telomerase/genética , Transcrição Gênica
17.
Proc Natl Acad Sci U S A ; 105(41): 15878-83, 2008 Oct 14.
Artigo em Inglês | MEDLINE | ID: mdl-18840680

RESUMO

Aire induces ectopic expression of peripheral tissue antigens (PTAs) in thymic medullary epithelial cells, which promotes immunological tolerance. Beginning with a broad screen of histone peptides, we demonstrate that the mechanism by which this single factor controls the transcription of thousands of genes involves recognition of the amino-terminal tail of histone H3, but not of other histones, by one of Aire's plant homeodomain (PHD) fingers. Certain posttranslational modifications of H3 tails, notably dimethylation or trimethylation at H3K4, abrogated binding by Aire, whereas others were tolerated. Similar PHD finger-H3 tail-binding properties were recently reported for BRAF-histone deacetylase complex 80 and DNA methyltransferase 3L; sequence alignment, molecular modeling, and biochemical analyses showed these factors and Aire to have structure-function relationships in common. In addition, certain PHD1 mutations underlying the polyendocrine disorder autoimmune polyendocrinopathy-candidiases-ectodermaldystrophy compromised Aire recognition of H3. In vitro binding assays demonstrated direct physical interaction between Aire and nucleosomes, which was in part buttressed by its affinity to DNA. In vivo Aire interactions with chromosomal regions depleted of H3K4me3 were dependent on its H3 tail-binding activity, and this binding was necessary but not sufficient for the up-regulation of genes encoding PTAs. Thus, Aire's activity as a histone-binding module mediates the thymic display of PTAs that promotes self-tolerance and prevents organ-specific autoimmunity.


Assuntos
Histonas/metabolismo , Tolerância Imunológica , Domínios e Motivos de Interação entre Proteínas/imunologia , Fatores de Transcrição/metabolismo , Animais , Autoantígenos/genética , Autoimunidade , Cromatina/metabolismo , Camundongos , Especificidade de Órgãos/imunologia , Ligação Proteica , Fatores de Transcrição/imunologia , Regulação para Cima , Proteína AIRE
18.
medRxiv ; 2021 Jan 29.
Artigo em Inglês | MEDLINE | ID: mdl-33532787

RESUMO

Coronavirus Disease 2019 (COVID-19), caused by Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2), is associated with a wide range of clinical manifestations, including autoimmune features and autoantibody production. We developed three different protein arrays to measure hallmark IgG autoantibodies associated with Connective Tissue Diseases (CTDs), Anti-Cytokine Antibodies (ACA), and anti-viral antibody responses in 147 hospitalized COVID-19 patients in three different centers. Autoantibodies were identified in approximately 50% of patients, but in <15% of healthy controls. When present, autoantibodies largely targeted autoantigens associated with rare disorders such as myositis, systemic sclerosis and CTD overlap syndromes. Anti-nuclear antibodies (ANA) were observed in ∼25% of patients. Patients with autoantibodies tended to demonstrate one or a few specificities whereas ACA were even more prevalent, and patients often had antibodies to multiple cytokines. Rare patients were identified with IgG antibodies against angiotensin converting enzyme-2 (ACE-2). A subset of autoantibodies and ACA developed de novo following SARS-CoV-2 infection while others were transient. Autoantibodies tracked with longitudinal development of IgG antibodies that recognized SARS-CoV-2 structural proteins such as S1, S2, M, N and a subset of non-structural proteins, but not proteins from influenza, seasonal coronaviruses or other pathogenic viruses. COVID-19 patients with one or more autoantibodies tended to have higher levels of antibodies against SARS-CoV-2 Nonstructural Protein 1 (NSP1) and Methyltransferase (ME). We conclude that SARS-CoV-2 causes development of new-onset IgG autoantibodies in a significant proportion of hospitalized COVID-19 patients and are positively correlated with immune responses to SARS-CoV-2 proteins.

19.
Nat Commun ; 12(1): 5417, 2021 09 14.
Artigo em Inglês | MEDLINE | ID: mdl-34521836

RESUMO

COVID-19 is associated with a wide range of clinical manifestations, including autoimmune features and autoantibody production. Here we develop three protein arrays to measure IgG autoantibodies associated with connective tissue diseases, anti-cytokine antibodies, and anti-viral antibody responses in serum from 147 hospitalized COVID-19 patients. Autoantibodies are identified in approximately 50% of patients but in less than 15% of healthy controls. When present, autoantibodies largely target autoantigens associated with rare disorders such as myositis, systemic sclerosis and overlap syndromes. A subset of autoantibodies targeting traditional autoantigens or cytokines develop de novo following SARS-CoV-2 infection. Autoantibodies track with longitudinal development of IgG antibodies recognizing SARS-CoV-2 structural proteins and a subset of non-structural proteins, but not proteins from influenza, seasonal coronaviruses or other pathogenic viruses. We conclude that SARS-CoV-2 causes development of new-onset IgG autoantibodies in a significant proportion of hospitalized COVID-19 patients and are positively correlated with immune responses to SARS-CoV-2 proteins.


Assuntos
Autoanticorpos/imunologia , COVID-19/imunologia , Imunoglobulina G/imunologia , SARS-CoV-2/imunologia , Idoso , Anticorpos Antinucleares/sangue , Anticorpos Antinucleares/imunologia , Anticorpos Antivirais/sangue , Anticorpos Antivirais/imunologia , Autoanticorpos/sangue , Autoantígenos/imunologia , Doenças do Tecido Conjuntivo/imunologia , Citocinas/imunologia , Feminino , Hospitalização , Humanos , Imunoglobulina G/sangue , Masculino , Pessoa de Meia-Idade , SARS-CoV-2/patogenicidade , Proteínas Virais/imunologia
20.
J Biol Chem ; 284(49): 34283-95, 2009 Dec 04.
Artigo em Inglês | MEDLINE | ID: mdl-19808676

RESUMO

The NSD (nuclear receptor SET domain-containing) family of histone lysine methyltransferases is a critical participant in chromatin integrity as evidenced by the number of human diseases associated with the aberrant expression of its family members. Yet, the specific targets of these enzymes are not clear, with marked discrepancies being reported in the literature. We demonstrate that NSD2 can exhibit disparate target preferences based on the nature of the substrate provided. The NSD2 complex purified from human cells and recombinant NSD2 both exhibit specific targeting of histone H3 lysine 36 (H3K36) when provided with nucleosome substrates, but histone H4 lysine 44 is the primary target in the case of octamer substrates, irrespective of the histones being native or recombinant. This disparity is negated when NSD2 is presented with octamer targets in conjunction with short single- or double-stranded DNA. Although the octamers cannot form nucleosomes, the target is nonetheless nucleosome-specific as is the product, dimethylated H3K36. This study clarifies in part the previous discrepancies reported with respect to NSD targets. We propose that DNA acts as an allosteric effector of NSD2 such that H3K36 becomes the preferred target.


Assuntos
Histona-Lisina N-Metiltransferase/química , Animais , Linhagem Celular Tumoral , Cromatina/química , DNA/química , Vetores Genéticos , Histona-Lisina N-Metiltransferase/metabolismo , Histonas/química , Humanos , Lisina/química , Espectrometria de Massas/métodos , Nucleossomos/química , Processamento de Proteína Pós-Traducional , Estrutura Terciária de Proteína , Proteínas Recombinantes/química , Xenopus laevis
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