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1.
PLoS Pathog ; 16(6): e1008555, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32579593

RESUMO

Exhaustion is a dysfunctional state of cytotoxic CD8+ T cells (CTL) observed in chronic infection and cancer. Current in vivo models of CTL exhaustion using chronic viral infections or cancer yield very few exhausted CTL, limiting the analysis that can be done on these cells. Establishing an in vitro system that rapidly induces CTL exhaustion would therefore greatly facilitate the study of this phenotype, identify the truly exhaustion-associated changes and allow the testing of novel approaches to reverse or prevent exhaustion. Here we show that repeat stimulation of purified TCR transgenic OT-I CTL with their specific peptide induces all the functional (reduced cytokine production and polyfunctionality, decreased in vivo expansion capacity) and phenotypic (increased inhibitory receptors expression and transcription factor changes) characteristics of exhaustion. Importantly, in vitro exhausted cells shared the transcriptomic characteristics of the gold standard of exhaustion, CTL from LCMV cl13 infections. Gene expression of both in vitro and in vivo exhausted CTL was distinct from T cell anergy. Using this system, we show that Tcf7 promoter DNA methylation contributes to TCF1 downregulation in exhausted CTL. Thus this novel in vitro system can be used to identify genes and signaling pathways involved in exhaustion and will facilitate the screening of reagents that prevent/reverse CTL exhaustion.


Assuntos
Linfócitos T CD8-Positivos/imunologia , Metilação de DNA/imunologia , Fator 1-alfa Nuclear de Hepatócito/imunologia , Coriomeningite Linfocítica/imunologia , Vírus da Coriomeningite Linfocítica/imunologia , Regiões Promotoras Genéticas/imunologia , Animais , Linfócitos T CD8-Positivos/patologia , Fator 1-alfa Nuclear de Hepatócito/genética , Coriomeningite Linfocítica/genética , Coriomeningite Linfocítica/patologia , Vírus da Coriomeningite Linfocítica/genética , Camundongos , Camundongos Transgênicos , Transdução de Sinais/genética , Transdução de Sinais/imunologia
2.
Immunity ; 52(5): 825-841.e8, 2020 05 19.
Artigo em Inglês | MEDLINE | ID: mdl-32396847

RESUMO

CD8+ T cell exhaustion is a major barrier to current anti-cancer immunotherapies. Despite this, the developmental biology of exhausted CD8+ T cells (Tex) remains poorly defined, restraining improvement of strategies aimed at "re-invigorating" Tex cells. Here, we defined a four-cell-stage developmental framework for Tex cells. Two TCF1+ progenitor subsets were identified, one tissue restricted and quiescent and one more blood accessible, that gradually lost TCF1 as it divided and converted to a third intermediate Tex subset. This intermediate subset re-engaged some effector biology and increased upon PD-L1 blockade but ultimately converted into a fourth, terminally exhausted subset. By using transcriptional and epigenetic analyses, we identified the control mechanisms underlying subset transitions and defined a key interplay between TCF1, T-bet, and Tox in the process. These data reveal a four-stage developmental hierarchy for Tex cells and define the molecular, transcriptional, and epigenetic mechanisms that could provide opportunities to improve cancer immunotherapy.


Assuntos
Linfócitos T CD8-Positivos/imunologia , Epigênese Genética/imunologia , Neoplasias/imunologia , Subpopulações de Linfócitos T/imunologia , Transcrição Genética/imunologia , Animais , Antígeno B7-H1/genética , Antígeno B7-H1/imunologia , Linfócitos T CD8-Positivos/citologia , Linfócitos T CD8-Positivos/metabolismo , Células Cultivadas , Epigênese Genética/genética , Fator 1-alfa Nuclear de Hepatócito/genética , Fator 1-alfa Nuclear de Hepatócito/imunologia , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/imunologia , Humanos , Imunoterapia/métodos , Camundongos Endogâmicos C57BL , Neoplasias/genética , Neoplasias/terapia , Proteínas com Domínio T/genética , Proteínas com Domínio T/imunologia , Subpopulações de Linfócitos T/metabolismo , Transcrição Genética/genética
3.
J Immunol ; 203(4): 801-806, 2019 08 15.
Artigo em Inglês | MEDLINE | ID: mdl-31300510

RESUMO

Differentiation of T follicular helper (TFH) cells is regulated by a complex transcriptional network, with mutually antagonistic Bcl6-Blimp1 as a core regulatory axis. It is well established that Tcf1 acts upstream of Bcl6 for its optimal induction to program TFH cell differentiation. In this study, we show that whereas genetic ablation of Tcf1 in mice greatly diminished TFH cells in response to viral infection, compound deletion of Blimp1 with Tcf1 restored TFH cell frequency, numbers, and generation of germinal center B cells. Aberrant upregulation of T-bet and Id2 in Tcf1-deficient TFH cells was also largely rectified by ablating Blimp1. Tcf1 chromatin immunoprecipitation sequencing in TFH cells identified two strong Tcf1 binding sites in the Blimp1 gene at a 24-kb upstream and an intron-3 element. Deletion of the intron-3 element, but not the 24-kb upstream element, compromised production of TFH cells. Our data demonstrate that Tcf1-mediated Blimp1 repression is functionally critical for safeguarding TFH cell differentiation.


Assuntos
Diferenciação Celular/imunologia , Fator 1-alfa Nuclear de Hepatócito/imunologia , Fator 1 de Ligação ao Domínio I Regulador Positivo/imunologia , Linfócitos T Auxiliares-Indutores/citologia , Viroses/imunologia , Animais , Regulação da Expressão Gênica/imunologia , Centro Germinativo/imunologia , Fator 1-alfa Nuclear de Hepatócito/metabolismo , Ativação Linfocitária/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Mutantes , Fator 1 de Ligação ao Domínio I Regulador Positivo/metabolismo , Linfócitos T Auxiliares-Indutores/imunologia , Linfócitos T Auxiliares-Indutores/metabolismo
4.
Nat Immunol ; 20(9): 1150-1160, 2019 09.
Artigo em Inglês | MEDLINE | ID: mdl-31358996

RESUMO

Innate lymphoid cells (ILCs) play important functions in immunity and tissue homeostasis, but their development is poorly understood. Through the use of single-cell approaches, we examined the transcriptional and functional heterogeneity of ILC progenitors, and studied the precursor-product relationships that link the subsets identified. This analysis identified two successive stages of ILC development within T cell factor 1-positive (TCF-1+) early innate lymphoid progenitors (EILPs), which we named 'specified EILPs' and 'committed EILPs'. Specified EILPs generated dendritic cells, whereas this potential was greatly decreased in committed EILPs. TCF-1 was dispensable for the generation of specified EILPs, but required for the generation of committed EILPs. TCF-1 used a pre-existing regulatory landscape established in upstream lymphoid precursors to bind chromatin in EILPs. Our results provide insight into the mechanisms by which TCF-1 promotes developmental progression of ILC precursors, while constraining their dendritic cell lineage potential and enforcing commitment to ILC fate.


Assuntos
Linhagem da Célula/imunologia , Células Dendríticas/citologia , Fator 1-alfa Nuclear de Hepatócito/imunologia , Células Progenitoras Linfoides/citologia , Linfócitos T/citologia , Animais , Diferenciação Celular/imunologia , Células Cultivadas , Regulação da Expressão Gênica/genética , Fator 1-alfa Nuclear de Hepatócito/genética , Camundongos , Camundongos Endogâmicos C57BL , Transcrição Genética/genética
5.
J Immunol ; 203(2): 323-327, 2019 07 15.
Artigo em Inglês | MEDLINE | ID: mdl-31175159

RESUMO

The differentiation of memory CD8+ T cells is critical to the long-term cellular immunity. The transcription factor BCL6 has been reportedly important for the generation and maintenance of memory CD8+ T cells; however, using the newly established BCL6 conditional knockout mouse model, we demonstrate that BCL6 is dispensable for the maintenance of established memory CD8+ T cell pool, although BCL6 is still required for the generation of CD8+ memory precursors upon acute viral infection. In addition, BCL6 promotes the expression of TCF-1 via directly binding to the Tcf7 (gene symbol for TCF-1) allele in CD8+ memory precursors and forced expression of TCF-1 restores the generation of BCL6-deficient memory precursors. Thus, our findings clarify that BCL6 is dispensable for the maintenance of memory CD8+ T cells, but functions as an important upstream of TCF-1 to regulate the generation of memory precursors in acute viral infection.


Assuntos
Linfócitos T CD8-Positivos/imunologia , Memória Imunológica/genética , Proteínas Proto-Oncogênicas c-bcl-6/genética , Fatores de Transcrição/genética , Viroses/genética , Doença Aguda , Animais , Diferenciação Celular/genética , Diferenciação Celular/imunologia , Fator 1-alfa Nuclear de Hepatócito/genética , Fator 1-alfa Nuclear de Hepatócito/imunologia , Memória Imunológica/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Proteínas Proto-Oncogênicas c-bcl-6/imunologia , Fatores de Transcrição/imunologia , Viroses/imunologia
6.
J Immunol ; 202(8): 2296-2306, 2019 04 15.
Artigo em Inglês | MEDLINE | ID: mdl-30814306

RESUMO

NK cells are innate-like lymphocytes that eliminate virally infected and cancerous cells, but the mechanisms that control NK cell development and cytotoxicity are incompletely understood. We identified roles for sclerostin domain-containing-1 (Sostdc1) in NK cell development and function. Sostdc1-knockout (Sostdc1 -/-) mice display a progressive accumulation of transitional NK cells (tNKs) (CD27+CD11b+) with age, indicating a partial developmental block. The NK cell Ly49 repertoire in Sostdc1 -/- mice is also changed. Lower frequencies of Sostdc1 -/- splenic tNKs express inhibitory Ly49G2 receptors, but higher frequencies express activating Ly49H and Ly49D receptors. However, the frequencies of Ly49I+, G2+, H+, and D+ populations were universally decreased at the most mature (CD27-CD11b+) stage. We hypothesized that the Ly49 repertoire in Sostdc1 -/- mice would correlate with NK killing ability and observed that Sostdc1-/- NK cells are hyporesponsive against MHC class I-deficient cell targets in vitro and in vivo, despite higher CD107a surface levels and similar IFN-γ expression to controls. Consistent with Sostdc1's known role in Wnt signaling regulation, Tcf7 and Lef1 levels were higher in Sostdc1 -/- NK cells. Expression of the NK development gene Id2 was decreased in Sostdc1-/- immature NK and tNK cells, but Eomes and Tbx21 expression was unaffected. Reciprocal bone marrow transplant experiments showed that Sostdc1 regulates NK cell maturation and expression of Ly49 receptors in a cell-extrinsic fashion from both nonhematopoietic and hematopoietic sources. Taken together, these data support a role for Sostdc1 in the regulation of NK cell maturation and cytotoxicity, and identify potential NK cell niches.


Assuntos
Proteínas Morfogenéticas Ósseas/imunologia , Imunidade Celular , Células Matadoras Naturais/imunologia , Via de Sinalização Wnt/imunologia , Proteínas Adaptadoras de Transdução de Sinal , Animais , Proteínas Morfogenéticas Ósseas/genética , Fator 1-alfa Nuclear de Hepatócito/genética , Fator 1-alfa Nuclear de Hepatócito/imunologia , Proteína 2 Inibidora de Diferenciação/genética , Proteína 2 Inibidora de Diferenciação/imunologia , Células Matadoras Naturais/citologia , Fator 1 de Ligação ao Facilitador Linfoide/genética , Fator 1 de Ligação ao Facilitador Linfoide/imunologia , Camundongos , Camundongos Knockout , Proteínas com Domínio T/genética , Proteínas com Domínio T/imunologia , Via de Sinalização Wnt/genética
7.
Nat Immunol ; 19(12): 1366-1378, 2018 12.
Artigo em Inglês | MEDLINE | ID: mdl-30420627

RESUMO

Thymocyte development requires a complex orchestration of multiple transcription factors. Ablating either TCF-1 or HEB in CD4+CD8+ thymocytes elicits similar developmental outcomes including increased proliferation, decreased survival, and fewer late Tcra rearrangements. Here, we provide a mechanistic explanation for these similarities by showing that TCF-1 and HEB share ~7,000 DNA-binding sites genome wide and promote chromatin accessibility. The binding of both TCF-1 and HEB was required at these shared sites for epigenetic and transcriptional gene regulation. Binding of TCF-1 and HEB to their conserved motifs in the enhancer regions of genes associated with T cell differentiation promoted their expression. Binding to sites lacking conserved motifs in the promoter regions of cell-cycle-associated genes limited proliferation. TCF-1 displaced nucleosomes, allowing for chromatin accessibility. Importantly, TCF-1 inhibited Notch signaling and consequently protected HEB from Notch-mediated proteasomal degradation. Thus, TCF-1 shifts nucleosomes and safeguards HEB, thereby enabling their cooperation in establishing the epigenetic and transcription profiles of CD4+CD8+ thymocytes.


Assuntos
Fatores de Transcrição Hélice-Alça-Hélice Básicos/imunologia , Regulação da Expressão Gênica/imunologia , Fator 1-alfa Nuclear de Hepatócito/imunologia , Linfopoese/imunologia , Timócitos/imunologia , Animais , Antígenos CD4/imunologia , Antígenos CD8/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos
8.
Blood Adv ; 2(14): 1685-1690, 2018 07 24.
Artigo em Inglês | MEDLINE | ID: mdl-30021780

RESUMO

Expression of the transcription factor T-cell factor 1 (TCF1) identifies antigen-experienced murine CD8+ T cells that retain potential for lymphoid recirculation and the ability to self-renew while producing more differentiated effector cells. We found that CD8+ T cells in the blood of both healthy and chronically infected humans expressed TCF1 at 3 distinct levels: high (TCF1-hi), intermediate (TCF1-int), and low (TCF1-lo). TCF1-hi cells could be found within both the naive and memory compartments and were characterized by relative quiescence and lack of immediate effector function. A substantial fraction of TCF1-int cells were found among memory cells, and TCF1-int cells exhibited robust immediate effector functions. TCF1-lo cells were most enriched in effector memory cells that expressed the senescence marker CD57. Following reactivation, TCF1-hi cells gave rise to TCF1-lo descendants while self-renewing the TCF1-hi progenitor. By contrast, reactivation of TCF1-lo cells produced more TCF1-lo cells without evidence of de-differentiating into TCF1-hi cells. Flow cytometric analyses of TCF1 expression from patient specimens may become a useful biomarker for adaptive immune function in response to vaccination, infection, autoimmunity, and cancer.


Assuntos
Linfócitos T CD8-Positivos/metabolismo , Regulação da Expressão Gênica , Fator 1-alfa Nuclear de Hepatócito/biossíntese , Memória Imunológica , Linfócitos T CD8-Positivos/citologia , Linfócitos T CD8-Positivos/imunologia , Feminino , Fator 1-alfa Nuclear de Hepatócito/imunologia , Humanos , Masculino
9.
Biochem Med (Zagreb) ; 28(2): 020703, 2018 Jun 15.
Artigo em Inglês | MEDLINE | ID: mdl-29666556

RESUMO

Introduction: Maturity onset diabetes of the young due to HNF1A mutations (HNF1A-MODY) is the most frequent form of monogenic diabetes in adults. It is often misdiagnosed as type 1 or type 2 diabetes, but establishing genetic diagnosis is important, as treatment differs from the common types of diabetes. HNF1A-MODY has not been investigated in Croatia before due to limited access to genetic testing. In this study we aimed to describe the characteristics of young adults diagnosed with diabetes before the age of 45 years, who have rare HNF1A allele variants, and estimate the prevalence of HNF1A-MODY in Croatia. Materials and methods: We recruited 477 C-peptide positive and beta cell antibody negative subjects through the Croatian Diabetes Registry. HNF1A was sequenced for all participants and systematic assessment of the variants found was performed. The prevalence of HNF1A-MODY was calculated in the study group and results extrapolated to estimate the proportion of diabetic individuals with HNF1A-MODY in Croatia and the population prevalence. Results: Our study identified 13 individuals harbouring rare HNF1A allelic variants. After systematic assessment, 8 were assigned a diagnosis of HNF1A-MODY. Two individuals were able to discontinue insulin treatment following the diagnosis. We estimated that HNF1A-MODY in Croatia has a prevalence of 66 (95% CI 61 - 72) cases per million. Conclusions: The estimated prevalence of HNF1A-MODY in Croatia is similar to that reported in other European countries. Finding cases lead to important treatment changes for patients. This strongly supports the introduction of diagnostic genetic testing for monogenic diabetes in Croatia.


Assuntos
Peptídeo C/sangue , Diabetes Mellitus Tipo 2/diagnóstico , Diabetes Mellitus Tipo 2/epidemiologia , Fator 1-alfa Nuclear de Hepatócito/genética , Mutação , Sistema de Registros , Adolescente , Adulto , Idoso , Alelos , Autoanticorpos/sangue , Biomarcadores/sangue , Croácia/epidemiologia , Diabetes Mellitus Tipo 2/sangue , Diabetes Mellitus Tipo 2/genética , Feminino , Expressão Gênica , Frequência do Gene , Testes Genéticos , Fator 1-alfa Nuclear de Hepatócito/imunologia , Humanos , Células Secretoras de Insulina/imunologia , Células Secretoras de Insulina/patologia , Masculino , Pessoa de Meia-Idade , Prevalência , Análise de Sequência de DNA
10.
J Immunol ; 200(10): 3397-3406, 2018 05 15.
Artigo em Inglês | MEDLINE | ID: mdl-29632143

RESUMO

T cell factor 1 (TCF-1) is expressed in both developing and mature T cells and has been shown to restrain mature T cell-mediated Th17 responses by inhibiting IL-17 expression. However, it is not clear when TCF-1 is required in vivo to restrain the magnitude of peripheral Th17 responses and what the molecular mechanisms responsible for TCF-1-regulated IL-17 gene expression are. In this study, we showed that conditional deletion of TCF-1 at the early but not later CD4+CD8+ double-positive stage in mice enhanced Th17 differentiation and aggravated experimental autoimmune encephalomyelitis, which correlates with abnormally high IL-17 expression. Expression of TCF-1 in TCF-1-deficient thymocytes but not TCF-1-deficient Th17 cells inhibited IL-17 expression. TCF-1 binds to IL-17 promoter regions, and deletion of two TCF-1 binding sites relieves TCF-1-mediated inhibition of IL-17 promoter activity. Lastly, wild-type TCF-1, but not a TCF-1 mutant that has no intrinsic histone deacetylase activity, was able to inhibit IL-17 expression in TCF-1 deficient mouse thymocytes. Thus, our study demonstrates the requirement of TCF-1 in vivo at stages earlier than double-positive cells to restrain peripheral Th17 immunity by directly binding and inhibiting IL-17 promoter in its intrinsic histone deacetylase-dependent manner.


Assuntos
Expressão Gênica/imunologia , Fator 1-alfa Nuclear de Hepatócito/imunologia , Imunidade/genética , Imunidade/imunologia , Interleucina-17/genética , Interleucina-17/imunologia , Células Th17/imunologia , Animais , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD8-Positivos/imunologia , Diferenciação Celular/genética , Diferenciação Celular/imunologia , Encefalomielite Autoimune Experimental/genética , Encefalomielite Autoimune Experimental/imunologia , Expressão Gênica/genética , Histona Desacetilases/genética , Histona Desacetilases/imunologia , Camundongos , Regiões Promotoras Genéticas/genética , Regiões Promotoras Genéticas/imunologia , Timócitos/imunologia
11.
J Exp Med ; 215(2): 575-594, 2018 02 05.
Artigo em Inglês | MEDLINE | ID: mdl-29282254

RESUMO

Upon infection with an intracellular pathogen, cytotoxic CD8+ T cells develop diverse differentiation states characterized by function, localization, longevity, and the capacity for self-renewal. The program of differentiation is determined, in part, by FOXO1, a transcription factor known to integrate extrinsic input in order to specify survival, DNA repair, self-renewal, and proliferation. At issue is whether the state of T cell differentiation is specified by initial conditions of activation or is actively maintained. To study the spectrum of T cell differentiation, we have analyzed an infection with mouse cytomegalovirus, a persistent-latent virus that elicits different cytotoxic T cell responses characterized as acute resolving or inflationary. Our results show that FOXO1 is continuously required for all the phenotypic characteristics of memory-effector T cells such that with acute inactivation of the gene encoding FOXO1, T cells revert to a short-lived effector phenotype, exhibit reduced viability, and manifest characteristics of anergy.


Assuntos
Linfócitos T CD8-Positivos/imunologia , Anergia Clonal , Proteína Forkhead Box O1/imunologia , Memória Imunológica , Transferência Adotiva , Animais , Antígenos Virais , Linfócitos T CD8-Positivos/citologia , Diferenciação Celular/imunologia , Sobrevivência Celular/imunologia , Proteína Forkhead Box O1/deficiência , Proteína Forkhead Box O1/genética , Fator 1-alfa Nuclear de Hepatócito/imunologia , Lectinas Tipo C , Ativação Linfocitária , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Muromegalovirus/imunologia , Receptores Imunológicos/imunologia
12.
Immunity ; 47(6): 1129-1141.e5, 2017 12 19.
Artigo em Inglês | MEDLINE | ID: mdl-29246443

RESUMO

During chronic stimulation, CD8+ T cells acquire an exhausted phenotype characterized by expression of inhibitory receptors, down-modulation of effector function, and metabolic impairments. T cell exhaustion protects from excessive immunopathology but limits clearance of virus-infected or tumor cells. We transcriptionally profiled antigen-specific T cells from mice infected with lymphocytic choriomeningitis virus strains that cause acute or chronic disease. T cell exhaustion during chronic infection was driven by high amounts of T cell receptor (TCR)-induced transcription factors IRF4, BATF, and NFATc1. These regulators promoted expression of inhibitory receptors, including PD-1, and mediated impaired cellular metabolism. Furthermore, they repressed the expression of TCF1, a transcription factor required for memory T cell differentiation. Reducing IRF4 expression restored the functional and metabolic properties of antigen-specific T cells and promoted memory-like T cell development. These findings indicate that IRF4 functions as a central node in a TCR-responsive transcriptional circuit that establishes and sustains T cell exhaustion during chronic infection.


Assuntos
Linfócitos T CD8-Positivos/imunologia , Memória Imunológica , Fatores Reguladores de Interferon/imunologia , Coriomeningite Linfocítica/imunologia , Vírus da Coriomeningite Linfocítica/imunologia , Receptores de Antígenos de Linfócitos T/imunologia , Animais , Fatores de Transcrição de Zíper de Leucina Básica/genética , Fatores de Transcrição de Zíper de Leucina Básica/imunologia , Linfócitos T CD8-Positivos/virologia , Diferenciação Celular , Regulação da Expressão Gênica , Células HEK293 , Fator 1-alfa Nuclear de Hepatócito/genética , Fator 1-alfa Nuclear de Hepatócito/imunologia , Humanos , Fatores Reguladores de Interferon/deficiência , Fatores Reguladores de Interferon/genética , Ativação Linfocitária , Depleção Linfocítica , Coriomeningite Linfocítica/genética , Coriomeningite Linfocítica/virologia , Vírus da Coriomeningite Linfocítica/crescimento & desenvolvimento , Camundongos , Camundongos Knockout , Fatores de Transcrição NFATC/genética , Fatores de Transcrição NFATC/imunologia , Receptor de Morte Celular Programada 1/genética , Receptor de Morte Celular Programada 1/imunologia , Receptores de Antígenos de Linfócitos T/genética , Transdução de Sinais
13.
Proc Natl Acad Sci U S A ; 114(42): E8865-E8874, 2017 10 17.
Artigo em Inglês | MEDLINE | ID: mdl-28973925

RESUMO

The factors and steps controlling postinfection CD8+ T cell terminal effector versus memory differentiation are incompletely understood. Whereas we found that naive TCF7 (alias "Tcf-1") expression is FOXO1 independent, early postinfection we report bimodal, FOXO1-dependent expression of the memory-essential transcription factor TCF7 in pathogen-specific CD8+ T cells. We determined the early postinfection TCF7high population is marked by low TIM3 expression and bears memory signature hallmarks before the appearance of established memory precursor marker CD127 (IL-7R). These cells exhibit diminished TBET, GZMB, mTOR signaling, and cell cycle progression. Day 5 postinfection, TCF7high cells express higher memory-associated BCL2 and EOMES, as well as increased accumulation potential and capacity to differentiate into memory phenotype cells. TCF7 retroviral transduction opposes GZMB expression and the formation of KLRG1pos phenotype cells, demonstrating an active role for TCF7 in extinguishing the effector program and forestalling terminal differentiation. Past the peak of the cellular immune response, we report a gradient of FOXO1 and TCF7 expression, which functions to oppose TBET and orchestrate a continuum of effector-to-memory phenotypes.


Assuntos
Linfócitos T CD8-Positivos/fisiologia , Proteína Forkhead Box O1/metabolismo , Memória Imunológica/fisiologia , Animais , Infecções por Arenaviridae/imunologia , Infecções por Arenaviridae/patologia , Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/virologia , Diferenciação Celular , Proteína Forkhead Box O1/genética , Granzimas/genética , Granzimas/metabolismo , Receptor Celular 2 do Vírus da Hepatite A/genética , Receptor Celular 2 do Vírus da Hepatite A/metabolismo , Fator 1-alfa Nuclear de Hepatócito/genética , Fator 1-alfa Nuclear de Hepatócito/imunologia , Fator 1-alfa Nuclear de Hepatócito/metabolismo , Subunidade alfa de Receptor de Interleucina-7/imunologia , Subunidade alfa de Receptor de Interleucina-7/metabolismo , Lectinas Tipo C , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Fenótipo , Receptores Imunológicos/metabolismo
14.
Cell Rep ; 20(3): 613-626, 2017 07 18.
Artigo em Inglês | MEDLINE | ID: mdl-28723565

RESUMO

The transcription factor Tcf1 is essential for the development of natural killer (NK) cells. However, its precise role has not been clarified. Our combined analysis of Tcf1-deficient and transgenic mice indicated that Tcf1 guides NK cells through three stages of development. Tcf1 expression directed bone marrow progenitors toward the NK cell lineage and ensured the survival of NK-committed cells, and its downregulation was needed for terminal maturation. Impaired survival of NK-committed cells was due to excessive expression of granzyme B (GzmB) and other granzyme family members, which induced NK cell self-destruction during maturation and following activation with cytokines or target cells. Mechanistically, Tcf1 binding reduced the activity of a Gzmb-associated regulatory element, and this accounted for the reduced Gzmb expression in Tcf1-expressing NK cells. These data identify an unexpected requirement to limit the expression of cytotoxic effector molecules for the normal expansion and function of NK cells.


Assuntos
Regulação Enzimológica da Expressão Gênica/imunologia , Granzimas/imunologia , Fator 1-alfa Nuclear de Hepatócito/imunologia , Células Matadoras Naturais/imunologia , Animais , Granzimas/genética , Fator 1-alfa Nuclear de Hepatócito/genética , Camundongos , Camundongos Knockout
15.
Nat Commun ; 8: 15050, 2017 05 03.
Artigo em Inglês | MEDLINE | ID: mdl-28466857

RESUMO

Differentiation and fate of virus-specific CD8+ T cells after cessation of chronic antigen stimulation is unclear. Here we show that a TCF1+CD127+PD1+ hepatitis C virus (HCV)-specific CD8+ T-cell subset exists in chronically infected patients with phenotypic features of T-cell exhaustion and memory, both before and after treatment with direct acting antiviral (DAA) agents. This subset is maintained during, and for a long duration after, HCV elimination. After antigen re-challenge the less differentiated TCF1+CD127+PD1+ population expands, which is accompanied by emergence of terminally exhausted TCF1-CD127-PD1hi HCV-specific CD8+ T cells. These results suggest the TCF1+CD127+PD1+ HCV-specific CD8+ T-cell subset has memory-like characteristics, including antigen-independent survival and recall proliferation. We thus provide evidence for the establishment of memory-like virus-specific CD8+ T cells in a clinically relevant setting of chronic viral infection and we uncover their fate after cessation of chronic antigen stimulation, implicating a potential strategy for antiviral immunotherapy.


Assuntos
Linfócitos T CD8-Positivos/imunologia , Hepacivirus/imunologia , Hepatite C Crônica/imunologia , Fator 1-alfa Nuclear de Hepatócito/imunologia , Adulto , Idoso , Antivirais/uso terapêutico , Linfócitos T CD8-Positivos/metabolismo , Linfócitos T CD8-Positivos/virologia , Feminino , Hepacivirus/efeitos dos fármacos , Hepacivirus/fisiologia , Hepatite C Crônica/metabolismo , Hepatite C Crônica/virologia , Fator 1-alfa Nuclear de Hepatócito/metabolismo , Humanos , Memória Imunológica/imunologia , Ativação Linfocitária/imunologia , Masculino , Pessoa de Meia-Idade , Subpopulações de Linfócitos T/imunologia , Subpopulações de Linfócitos T/metabolismo , Subpopulações de Linfócitos T/virologia , Adulto Jovem
16.
Cell Rep ; 17(2): 436-447, 2016 10 04.
Artigo em Inglês | MEDLINE | ID: mdl-27705792

RESUMO

Innate lymphoid cells (ILCs) are enriched at mucosal surfaces, where they provide immune surveillance. All ILC subsets develop from a common progenitor that gives rise to pre-committed progenitors for each of the ILC lineages. Currently, the temporal control of gene expression that guides the emergence of these progenitors is poorly understood. We used global transcriptional mapping to analyze gene expression in different ILC progenitors. We identified PD-1 to be specifically expressed in PLZF+ ILCp and revealed that the timing and order of expression of the transcription factors NFIL3, ID2, and TCF-1 was critical. Importantly, induction of ILC lineage commitment required only transient expression of NFIL3 prior to ID2 and TCF-1 expression. These findings highlight the importance of the temporal program that permits commitment of progenitors to the ILC lineage, and they expand our understanding of the core transcriptional program by identifying potential regulators of ILC development.


Assuntos
Fatores de Transcrição de Zíper de Leucina Básica/genética , Fator 1-alfa Nuclear de Hepatócito/genética , Imunidade Inata/imunologia , Linfócitos/imunologia , Receptor de Morte Celular Programada 1/genética , Animais , Fatores de Transcrição de Zíper de Leucina Básica/imunologia , Células da Medula Óssea/imunologia , Diferenciação Celular/genética , Diferenciação Celular/imunologia , Linhagem da Célula/imunologia , Regulação da Expressão Gênica , Fator 1-alfa Nuclear de Hepatócito/imunologia , Imunidade Inata/genética , Células Matadoras Naturais/imunologia , Camundongos , Receptor de Morte Celular Programada 1/imunologia , Fatores de Transcrição/genética , Fatores de Transcrição/imunologia
17.
Blood ; 128(4): 508-18, 2016 07 28.
Artigo em Inglês | MEDLINE | ID: mdl-27259979

RESUMO

Dendritic cells (DCs) are critical immune response regulators; however, the mechanism of DC differentiation is not fully understood. Heterozygous germ line GATA2 mutations induce GATA2-deficiency syndrome, characterized by monocytopenia, a predisposition to myelodysplasia/acute myeloid leukemia, and a profoundly reduced DC population, which is associated with increased susceptibility to viral infections, impaired phagocytosis, and decreased cytokine production. To define the role of GATA2 in DC differentiation and function, we studied Gata2 conditional knockout and haploinsufficient mice. Gata2 conditional deficiency significantly reduced the DC count, whereas Gata2 haploinsufficiency did not affect this population. GATA2 was required for the in vitro generation of DCs from Lin(-)Sca-1(+)Kit(+) cells, common myeloid-restricted progenitors, and common dendritic cell precursors, but not common lymphoid-restricted progenitors or granulocyte-macrophage progenitors, suggesting that GATA2 functions in the myeloid pathway of DC differentiation. Moreover, expression profiling demonstrated reduced expression of myeloid-related genes, including mafb, and increased expression of T-lymphocyte-related genes, including Gata3 and Tcf7, in Gata2-deficient DC progenitors. In addition, GATA2 was found to bind an enhancer element 190-kb downstream region of Gata3, and a reporter assay exhibited significantly reduced luciferase activity after adding this enhancer region to the Gata3 promoter, which was recovered by GATA sequence deletion within Gata3 +190. These results suggest that GATA2 plays an important role in cell-fate specification toward the myeloid vs T-lymphocyte lineage by regulating lineage-specific transcription factors in DC progenitors, thereby contributing to DC differentiation.


Assuntos
Diferenciação Celular/imunologia , Células Dendríticas/imunologia , Fator de Transcrição GATA2/imunologia , Animais , Diferenciação Celular/genética , Células Dendríticas/citologia , Fator de Transcrição GATA2/genética , Fator de Transcrição GATA3/genética , Fator de Transcrição GATA3/imunologia , Fator 1-alfa Nuclear de Hepatócito/genética , Fator 1-alfa Nuclear de Hepatócito/imunologia , Camundongos , Camundongos Knockout , Células Mieloides/citologia , Células Mieloides/imunologia , Linfócitos T/citologia , Linfócitos T/imunologia
18.
Nat Commun ; 7: 11171, 2016 Apr 06.
Artigo em Inglês | MEDLINE | ID: mdl-27048872

RESUMO

The gradual reprogramming of haematopoietic precursors into the T-cell fate is characterized by at least two sequential developmental stages. Following Notch1-dependent T-cell lineage specification during which the first T-cell lineage genes are expressed and myeloid and dendritic cell potential is lost, T-cell specific transcription factors subsequently induce T-cell commitment by repressing residual natural killer (NK)-cell potential. How these processes are regulated in human is poorly understood, especially since efficient T-cell lineage commitment requires a reduction in Notch signalling activity following T-cell specification. Here, we show that GATA3, in contrast to TCF1, controls human T-cell lineage commitment through direct regulation of three distinct processes: repression of NK-cell fate, upregulation of T-cell lineage genes to promote further differentiation and restraint of Notch activity. Repression of the Notch1 target gene DTX1 hereby is essential to prevent NK-cell differentiation. Thus, GATA3-mediated positive and negative feedback mechanisms control human T-cell lineage commitment.


Assuntos
Linhagem da Célula/genética , Retroalimentação Fisiológica , Fator de Transcrição GATA3/genética , Células-Tronco Hematopoéticas/imunologia , Timócitos/imunologia , Diferenciação Celular , Linhagem da Célula/imunologia , Reprogramação Celular , Criança , Fator de Transcrição GATA3/imunologia , Regulação da Expressão Gênica no Desenvolvimento , Células-Tronco Hematopoéticas/citologia , Fator 1-alfa Nuclear de Hepatócito/genética , Fator 1-alfa Nuclear de Hepatócito/imunologia , Humanos , Células Matadoras Naturais/citologia , Células Matadoras Naturais/imunologia , Cultura Primária de Células , Receptor Notch1/genética , Receptor Notch1/imunologia , Transdução de Sinais , Timócitos/citologia , Ubiquitina-Proteína Ligases/genética , Ubiquitina-Proteína Ligases/imunologia
19.
J Immunol ; 196(4): 1655-65, 2016 Feb 15.
Artigo em Inglês | MEDLINE | ID: mdl-26800876

RESUMO

The hepatocyte NF (HNF) family of transcription factors regulates the complex gene networks involved in lipid, carbohydrate, and protein metabolism. In humans, HNF1A mutations cause maturity onset of diabetes in the young type 3, whereas murine HNF6 participates in fetal liver B lymphopoiesis. In this study, we have identified a crucial role for the prototypical member of the family HNF1A in adult bone marrow B lymphopoiesis. HNF1A(-/-) mice exhibited a clear reduction in total blood and splenic B cells and a further pronounced one in transitional B cells. In HNF1A(-/-) bone marrow, all B cell progenitors-from pre-pro-/early pro-B cells to immature B cells-were dramatically reduced and their proliferation rate suppressed. IL-7 administration in vivo failed to boost B cell development in HNF1A(-/-) mice, whereas IL-7 stimulation of HNF1A(-/-) B cell progenitors in vitro revealed a marked impairment in STAT5 phosphorylation. The B cell differentiation potential of HNF1A(-/-) common lymphoid progenitors was severely impaired in vitro, and the expression of the B lymphopoiesis-promoting transcription factors E2A, EBF1, Pax5, and Bach2 was reduced in B cell progenitors in vivo. HNF1A(-/-) bone marrow chimera featured a dramatic defect in B lymphopoiesis recapitulating that of global HNF1A deficiency. The HNF1A(-/-) lymphopoiesis defect was confined to B cells as T lymphopoiesis was unaffected, and bone marrow common lymphoid progenitors and hematopoietic stem cells were even increased. Our data demonstrate that HNF1A is an important cell-intrinsic transcription factor in adult B lymphopoiesis and suggest the IL-7R/STAT5 module to be causally involved in mediating its function.


Assuntos
Linfócitos B/imunologia , Diferenciação Celular/imunologia , Fator 1-alfa Nuclear de Hepatócito/imunologia , Linfopoese/imunologia , Animais , Linfócitos B/citologia , Separação Celular , Citometria de Fluxo , Células Progenitoras Linfoides/citologia , Células Progenitoras Linfoides/imunologia , Camundongos , Camundongos Knockout , Reação em Cadeia da Polimerase em Tempo Real , Fatores de Transcrição
20.
Nat Immunol ; 17(3): 269-76, 2016 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-26779601

RESUMO

The precise lineage relationship between innate lymphoid cells (ILCs) and lymphoid tissue-inducer (LTi) cells is poorly understood. Using single-cell multiplex transcriptional analysis of 100 lymphoid genes and single-cell cultures of fetal liver precursor cells, we identified the common proximal precursor to these lineages and found that its bifurcation was marked by differential induction of the transcription factors PLZF and TCF1. Acquisition of individual effector programs specific to the ILC subsets ILC1, ILC2 and ILC3 was initiated later, at the common ILC precursor stage, by transient expression of mixed ILC1, ILC2 and ILC3 transcriptional patterns, whereas, in contrast, the development of LTi cells did not go through multilineage priming. Our findings provide insight into the divergent mechanisms of the differentiation of the ILC lineage and LTi cell lineage and establish a high-resolution 'blueprint' of their development.


Assuntos
Linhagem da Célula/imunologia , Subpopulações de Linfócitos/imunologia , Linfócitos/imunologia , Linfócitos T Auxiliares-Indutores/imunologia , Animais , Diferenciação Celular/genética , Citometria de Fluxo , Perfilação da Expressão Gênica , Fator 1-alfa Nuclear de Hepatócito/imunologia , Fatores de Transcrição Kruppel-Like/genética , Fatores de Transcrição Kruppel-Like/imunologia , Tecido Linfoide/citologia , Camundongos , Reação em Cadeia da Polimerase Multiplex , Proteína com Dedos de Zinco da Leucemia Promielocítica , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Análise de Célula Única
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