RESUMEN
The transcription factor Ets1 is essential for the development/differentiation of invariant Natural Killer T (iNKT) cells at multiple stages. However, its mechanisms of action and target genes in iNKT cells are still elusive. Here, we show that Ets1 is required for the optimal expression of the Vα14Jα18 T cell receptor (TCR) in post-selected thymic iNKT cells and their immediate differentiation. Ets1 is also critical for maintaining the peripheral homeostasis of iNKT cells, which is a role independent of the expression of the Vα14Jα18 TCR. Genome-wide transcriptomic analyses of post-selected iNKT cells further reveal that Ets1 controls leukocytes activation, proliferation differentiation, and leukocyte-mediated immunity. In addition, Ets1 regulates the expression of ICOS and PLZF in iNKT cells. More importantly, restoring the expression of PLZF and the Vα14Jα18 TCR partially rescues the differentiation of iNKT cells in the absence of Ets1. Taken together, our results establish a detailed molecular picture of how Ets1 regulates the stepwise differentiation of iNKT cells.
Asunto(s)
Diferenciación Celular/inmunología , Regulación de la Expresión Génica/inmunología , Células T Asesinas Naturales/inmunología , Proteína de la Leucemia Promielocítica con Dedos de Zinc/inmunología , Proteína Proto-Oncogénica c-ets-1/inmunología , Receptores de Antígenos de Linfocitos T alfa-beta/inmunología , Animales , Diferenciación Celular/genética , Ratones , Ratones Noqueados , Proteína de la Leucemia Promielocítica con Dedos de Zinc/genética , Proteína Proto-Oncogénica c-ets-1/genética , Receptores de Antígenos de Linfocitos T alfa-beta/genéticaRESUMEN
Establishment of immune tolerance is crucial to protect humans against asthma. Promyelocytic leukemia zinc finger (PLZF) is an emerging suppressor of inflammatory responses. CCL21-CCR7 signaling mediates tolerance development. However, whether PLZF and CCL21-CCR7 are required for the development of asthma tolerance is unknown. Here, we found that Zbtb16 (coding PLZF) and Ccl21 were upregulated in OVA-induced asthma tolerance (OT) lungs by RNA-seq. PLZF physically interacted with GATA3 and its expression was higher in GATA3+ Th2 cells and ILC2s in OT lungs. Zbtb16-knockdown in lymphocytes promoted the differentiation of CD3e+ CD4+ T cells, particularly those producing IL-4 and IL-5. Moreover, iNKT cells with high expression of PLZF were recruited into the lungs via draining lymph nodes during tolerance. Blockade of CCL21-CCR7 signaling in OT mice decreased the PLZF+ cell population, abolished CCR7-induced PLZF+ iNKT recruitment to the lungs, enhanced Th2responses and exacerbated lung pathology. In OT mice, respiratory syncytial virus (RSV) infection impeded PLZF+ cell and CCR7+ PLZF+ iNKT cellrecruitment to the lungs and increased airway resistance. Collectively, these results indicate that PLZF could interact with GATA3 and restrain differentiation of IL-4- and IL-5-producing T cells, iNKT cells with high PLZF expression are recruited to the lungs via CCL21-CCR7 signaling to facilitate the development of asthma tolerance.
Asunto(s)
Asma/inmunología , Quimiocina CCL21/inmunología , Tolerancia Inmunológica/inmunología , Pulmón/inmunología , Células T Asesinas Naturales/inmunología , Proteína de la Leucemia Promielocítica con Dedos de Zinc/inmunología , Receptores CCR7/inmunología , Animales , Linfocitos T CD4-Positivos/inmunología , Diferenciación Celular/inmunología , Línea Celular Tumoral , Modelos Animales de Enfermedad , Humanos , Masculino , Ratones , Ratones Endogámicos BALB C , Transducción de Señal/inmunología , Células Th2/inmunologíaRESUMEN
Following acute infection, herpes simplex virus 1 (HSV-1) lytic cycle viral gene expression is silenced; consequently, lifelong latency in neurons is established. Certain external stimuli that trigger reactivation from latency also activate the glucocorticoid receptor (GR). The synthetic corticosteroid dexamethasone, but not a GR-specific antagonist, increases the frequency of explant-induced reactivation from latency and stimulates productive infection. Furthermore, dexamethasone increases expression of cellular transcription factors in trigeminal ganglionic neurons: for example, SLUG and three Krüppel-like transcription factor (KLF) family members, KLF4, KLF15, and promyelocytic leukemia zinc finger protein (PLZF). Consequently, we hypothesized that stress-induced transcription factors stimulate expression of ICP4, a viral transcriptional regulator required for productive infection. New studies demonstrated that GR and KLF4, PLZF, or SLUG cooperatively transactivate the ICP4 enhancer upstream of a minimal promoter in monkey kidney cells (Vero) and mouse neuroblastoma cells (Neuro-2A). Strikingly, mutagenesis of two KLF4/Sp1 binding sites reduced GR- plus KLF4-, PLZF-, or SLUG-mediated transactivation to basal levels. A consensus enhancer (E)-Box adjacent to a KLF4/Sp1 binding site was also required for GR- and SLUG-, but not KLF family member-, mediated transactivation of the ICP4 promoter. Chromatin immunoprecipitation studies (ChIP) revealed GR and stress-induced transcription factors occupy ICP4 enhancer sequences. Conversely, specific binding was generally reduced in the KLF4/Sp1 mutant. Furthermore, GR and SLUG occupancy of ICP4 enhancer sequences was reduced in the E-Box mutant. Based on these studies, we suggest stressful stimuli can trigger productive infection because GR and specific stress-induced transcription factors activate ICP4 expression.IMPORTANCE Certain stressful stimuli activate the glucocorticoid receptor (GR) and increase the incidence of herpes simplex virus 1 (HSV-1) reactivation from latency. For example, a corticosteroid antagonist impairs productive infection and virus shedding following explant of trigeminal ganglia from latently infected mice. Infected cell protein 4 (ICP4) is the only immediate early viral transcriptional regulator required for productive infection, suggesting stressful stimuli stimulate ICP4 expression. New studies revealed GR and stress-induced transcription factors identified during reactivation from latency, SLUG and three Krüppel-like transcription factor family members (KLF4, KLF15, and promyelocytic leukemia zinc finger protein), cooperatively transactivate the ICP4 enhancer. Two KLF4 consensus binding sites were crucial for cooperative transactivation of the ICP4 enhancer. A consensus enhancer-box also mediated cooperative transactivation of the ICP4 enhancer by GR and SLUG. The ability of GR and stress-induced transcription factors to transactivate ICP4 enhancer activity is predicted to trigger productive infection following stressful stimuli.
Asunto(s)
Herpes Simple , Herpesvirus Humano 1/fisiología , Proteínas Inmediatas-Precoces/inmunología , Receptores de Glucocorticoides/inmunología , Activación Viral , Latencia del Virus , Animales , Línea Celular Tumoral , Chlorocebus aethiops , Regulación Viral de la Expresión Génica , Herpes Simple/inmunología , Herpes Simple/virología , Humanos , Factor 4 Similar a Kruppel , Factores de Transcripción de Tipo Kruppel/inmunología , Ratones , Proteína de la Leucemia Promielocítica con Dedos de Zinc/inmunología , Factores de Transcripción de la Familia Snail/inmunología , Activación Transcripcional , Células VeroRESUMEN
Innate lymphoid cells (ILCs) are generated early during ontogeny and persist predominantly as tissue-resident cells. Here, we examined how ILCs are maintained and renewed within tissues. We generated a single cell atlas of lung ILC2s and found that Il18r1+ ILCs comprise circulating and tissue-resident ILC progenitors (ILCP) and effector-cells with heterogeneous expression of the transcription factors Tcf7 and Zbtb16, and CD103. Our analyses revealed a continuous differentiation trajectory from Il18r1+ ST2- ILCPs to Il18r- ST2+ ILC2s, which was experimentally validated. Upon helminth infection, recruited and BM-derived cells generated the entire spectrum of ILC2s in parabiotic and shield chimeric mice, consistent with their potential role in the renewal of tissue ILC2s. Our findings identify local ILCPs and reveal ILCP in situ differentiation and tissue adaptation as a mechanism of ILC maintenance and phenotypic diversification. Local niches, rather than progenitor origin, or the developmental window during ontogeny, may dominantly imprint ILC phenotypes in adult tissues.
Asunto(s)
Inmunidad Innata/inmunología , Linfocitos/inmunología , Células Progenitoras Linfoides/inmunología , Animales , Diferenciación Celular/inmunología , Células Cultivadas , Femenino , Humanos , Subunidad alfa del Receptor de Interleucina-18/inmunología , Pulmón/inmunología , Ratones , Ratones Endogámicos C57BL , Proteína de la Leucemia Promielocítica con Dedos de Zinc/inmunología , Transducción de Señal/inmunología , Análisis de la Célula Individual/métodos , Factor 1 de Transcripción de Linfocitos T/inmunología , Factores de Transcripción/inmunologíaRESUMEN
The thymus generates cells of the T cell lineage that seed the lymphatic and blood systems. Transcription factor regulatory networks control the lineage programming and maturation of thymic precursor cells. Whether extrathymic antigenic events, such as the microbial colonization of the mucosal tract also shape the thymic T cell repertoire is unclear. We show here that intestinal microbes influence the thymic homeostasis of PLZF-expressing cells in early life. Impaired thymic development of PLZF+ innate lymphocytes in germ-free (GF) neonatal mice is restored by colonization with a human commensal, Bacteroides fragilis, but not with a polysaccharide A (PSA) deficient isogenic strain. Plasmacytoid dendritic cells influenced by microbes migrate from the colon to the thymus in early life to regulate PLZF+ cell homeostasis. Importantly, perturbations in thymic PLZF+ cells brought about by alterations in early gut microbiota persist into adulthood and are associated with increased susceptibility to experimental colitis. Our studies identify a pathway of communication between intestinal microbes and thymic lymphocytes in the neonatal period that can modulate host susceptibility to immune-mediated diseases later in life.
Asunto(s)
Microbioma Gastrointestinal , Linfocitos/inmunología , Timo/crecimiento & desarrollo , Animales , Bacterias/clasificación , Bacterias/genética , Bacterias/aislamiento & purificación , Bacteroides fragilis/fisiología , Diferenciación Celular , Colitis/genética , Colitis/inmunología , Colitis/microbiología , Colon/microbiología , Humanos , Linfocitos/citología , Ratones , Ratones Endogámicos C57BL , Proteína de la Leucemia Promielocítica con Dedos de Zinc/genética , Proteína de la Leucemia Promielocítica con Dedos de Zinc/inmunología , Timo/citología , Timo/inmunologíaRESUMEN
Human cytomegalovirus (HCMV) has been recently related with a lower susceptibility to multiple sclerosis (MS). HCMV promotes an adaptive development of NK cells bearing the CD94/NKG2C receptor with a characteristic phenotypic and functional profile. NK cells are proposed to play an immunoregulatory role in MS, and expansion of the NKG2C(+) subset was recently associated with reduced disability progression. To further explore this issue, additional adaptive NK cell markers, i.e., downregulation of FcεRIγ chain (FcRγ) and PLZF transcription factor, as well as antibody-dependent NK cell activation were assessed in controls and MS patients considering HCMV serology and clinical features. In line with previous reports, increased proportions of NKG2C(+), FcRγ(-), and PLZF(-) CD56dim NK cells were found in HCMV(+) cases. However, PLZF(-) NK cells were detected uncoupled from other adaptive markers within the CD56bright subset from HCMV(+) cases and among CD56dim NK cells from HCMV(-) MS patients, suggesting an additional effect of HCMV-independent factors in PLZF downregulation. Interferon-ß therapy was associated with lower proportions of FcRγ(-) CD56dim NK cells in HCMV(+) and increased PLZF(-) CD56bright NK cells in HCMV(-) patients, pointing out to an influence of the cytokine on the expression of adaptive NK cell-associated markers. In addition, proportions of NKG2C(+) and FcRγ(-) NK cells differed in progressive MS patients as compared to controls and other clinical forms. Remarkably, an adaptive NK cell phenotype did not directly correlate with enhanced antibody-triggered degranulation and TNFα production in MS in contrast to controls. Altogether, our results provide novel insights into the putative influence of HCMV and adaptive NK cells in MS.
Asunto(s)
Células Asesinas Naturales/inmunología , Activación de Linfocitos , Esclerosis Múltiple/inmunología , Adulto , Citomegalovirus/inmunología , Femenino , Humanos , Células Asesinas Naturales/patología , Masculino , Persona de Mediana Edad , Esclerosis Múltiple/patología , Subfamília C de Receptores Similares a Lectina de Células NK/inmunología , Subfamília D de Receptores Similares a Lectina de las Células NK/inmunología , Proteína de la Leucemia Promielocítica con Dedos de Zinc/inmunología , Estudios Prospectivos , Receptores Fc/inmunología , Factor de Necrosis Tumoral alfa/inmunologíaRESUMEN
Hepatic immune system is uniquely challenged to mount a controlled effector response to pathogens while maintaining tolerance to diet and microbial Ags. We have identified a novel population of innate-like, unconventional CD8αα+TCRαß+ T cells in naive mice and in human peripheral blood, called CD8αα Tunc, capable of controlling effector T cell responses. They are NK1.1+ (CD161+ in human), express NK-inhibitory receptors, and express the promyelocytic leukemia zinc finger (PLZF) transcription factor that distinguishes them from conventional CD8+ T cells. These cells display a cytotoxic phenotype and use a perforin-dependent mechanism to control Ag-induced or T cell-mediated autoimmune diseases. CD8αα Tunc are dependent upon IL-15/IL-2Rß signaling and PLZF for their development and/or survival. They are Foxp3-negative and their regulatory activity is associated with a functionally distinct Qa-1b-dependent population coexpressing CD11c and CD244. A polyclonal TCR repertoire, an activated/memory phenotype, and the presence of CD8αα Tunc in NKT- and in MAIT-deficient as well as in germ-free mice indicates that these cells recognize diverse self-protein Ags. Our studies reveal a distinct population of unconventional CD8+ T cells within the natural immune repertoire capable of controlling autoimmunity and also providing a new target for therapeutic intervention.
Asunto(s)
Autoinmunidad/inmunología , Linfocitos T CD8-positivos/inmunología , Hígado/inmunología , Proteína de la Leucemia Promielocítica con Dedos de Zinc/inmunología , Animales , Voluntarios Sanos , Humanos , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Ratones TransgénicosRESUMEN
Basophils are innate immune cells associated with type 2 immunity, allergic reactions, and host defense against parasite infections. In this study, we show that the transcription factor PLZF, which is known for its essential role in the function and development of several innate lymphocyte subsets, is also important for the myeloid-derived basophil lineage. PLZF-deficient mice had decreased numbers of basophil progenitors in the bone marrow and mature basophils in multiple peripheral tissues. Functionally, PLZF-deficient basophils were less responsive to IgE activation and produced reduced amounts of IL-4. The altered function of basophils resulted in a blunted Th2 T cell response to a protein allergen. Additionally, PLZF-deficient basophils had reduced expression of the IL-18 receptor, which impacted migration to lungs. PLZF, therefore, is a major player in controlling type 2 immune responses mediated not only by innate lymphocytes but also by myeloid-derived cells.
Asunto(s)
Basófilos/inmunología , Proteína de la Leucemia Promielocítica con Dedos de Zinc/inmunología , Factores de Transcripción/inmunología , Alérgenos/inmunología , Animales , Inmunidad Innata/inmunología , Inmunoglobulina E/inmunología , Interleucina-4/inmunología , Interleucina-8/inmunología , Subgrupos Linfocitarios/inmunología , Ratones , Ratones Noqueados , Células Mieloides/inmunología , Células Th2/inmunologíaRESUMEN
Pigs are important livestock and comprehensive understanding of their immune responses in infections is critical to improve vaccines and therapies. Moreover, similarities between human and swine physiology suggest that pigs are a superior animal model for immunological studies. However, paucity of experimental tools for a systematic analysis of the immune responses in pigs represent a major disadvantage. To evaluate the pig as a biomedical model and additionally expand the knowledge of rare immune cell populations in swine, we established a multicolor flow cytometry analysis platform of surface marker expression and cellular responses for porcine invariant Natural Killer T cells (iNKT). In humans, iNKT cells are among the first line defenders in various tissues, respond to CD1d-restricted antigens and become rapidly activated. Naïve porcine iNKT cells were CD3+/CD4-/CD8+ or CD3+/CD4-/CD8- and displayed an effector- or memory-like phenotype (CD25+/ICOS+/CD5hi/CD45RA-/CCR7 ± /CD27+). Based on their expression of the transcription factors T bet and the iNKT cell-specific promyelocytic leukemia zinc finger protein (PLZF), porcine iNKT cells were differentiated into functional subsets. Analogous to human iNKT cells, in vitro stimulation of porcine leukocytes with the CD1d ligand α-galactosylceramide resulted in rapid iNKT cell proliferation, evidenced by an increase in frequency and Ki-67 expression. Moreover, this approach revealed CD25, CD5, ICOS, and the major histocompatibility complex class II (MHC II) as activation markers on porcine iNKT cells. Activated iNKT cells also expressed interferon-γ, upregulated perforin expression, and displayed degranulation. In steady state, iNKT cell frequency was highest in newborn piglets and decreased with age. Upon infection with two viruses of high relevance to swine and humans, iNKT cells expanded. Animals infected with African swine fever virus displayed an increase of iNKT cell frequency in peripheral blood, regional lymph nodes, and lungs. During Influenza A virus infection, iNKT cell percentage increased in blood, lung lymph nodes, and broncho-alveolar lavage. Our in-depth characterization of porcine iNKT cells contributes to a better understanding of porcine immune responses, thereby facilitating the design of innovative interventions against infectious diseases. Moreover, we provide new evidence that endorses the suitability of the pig as a biomedical model for iNKT cell research.
Asunto(s)
Antígenos CD1d/inmunología , Antígenos CD/inmunología , Células T Asesinas Naturales/inmunología , Virosis/inmunología , Animales , Animales Recién Nacidos , Antígenos CD/metabolismo , Antígenos CD1d/metabolismo , Diferenciación Celular/inmunología , Proliferación Celular , Células Cultivadas , Citometría de Flujo/métodos , Galactosilceramidas/inmunología , Humanos , Inmunofenotipificación , Interferón gamma/inmunología , Interferón gamma/metabolismo , Leucocitos Mononucleares/citología , Leucocitos Mononucleares/inmunología , Leucocitos Mononucleares/metabolismo , Modelos Animales , Células T Asesinas Naturales/metabolismo , Proteína de la Leucemia Promielocítica con Dedos de Zinc/inmunología , Proteína de la Leucemia Promielocítica con Dedos de Zinc/metabolismo , Porcinos , Virosis/metabolismo , Virosis/virologíaRESUMEN
Natural killer T (NKT) cells are αß T cell receptor (TCR) expressing innate-like T cells that display natural killer (NK) cell markers. Based on TCR characteristics, they are divided into two groups restricted to the MHC class I-like molecule CD1d. Type I NKT cells, most extensively studied, are identified by a semi-invariant Vα14-Jα18 (mouse, Vα24-Jα18 in humans) TCR reactive to the prototypic ligand α-galactosylceramide presented on CD1d. In contrast, type II NKT cells display diverse TCR reacting to different CD1d-presented ligands. There are no reagents that identify all type II NKT cells, limiting their exploration. Here, we searched for novel type II NKT cells by comparing Jα18-/- MHCII-/- mice that harbour type II but not type I NKT cells, and CD1d-/- MHCII-/- mice, lacking all NKT cells. We identified significantly larger populations of CD4+ and CD4- CD8- (double negative, DN) TCRß+ cells expressing NKG2D or NKG2A/C/E in Jα18-/- MHCII-/- mice compared with CD1d-/- MHCII-/- mice, suggesting that 30%-50% of these cells were type II NKT cells. They expressed CD122, NK1.1, CXCR3 and intermediate/low levels of CD45RB. Further, the CD4+ subset was CD69+ , while the DN cells were CD49b+ and CD62L+ . Both subsets expressed the NKT cell-associated promyelocytic leukaemia zinc finger (PLZF) transcription factor and Tbet, while fewer cells expressed RORγt. NKG2D+ CD4+ and DN populations were producers of IFN-γ, but rarely IL-4 and IL-17. Taken together, we identify a novel subset of primary CD4+ and DN type II NKT cells that expresses NKG2 receptors have typical NKT cell phenotypes and a TH1-like cytokine production.
Asunto(s)
Antígenos CD1d/inmunología , Antígenos CD1d/metabolismo , Biomarcadores/metabolismo , Células Asesinas Naturales/inmunología , Células T Asesinas Naturales/inmunología , Animales , Femenino , Galactosilceramidas/inmunología , Galactosilceramidas/metabolismo , Interferón gamma/inmunología , Interferón gamma/metabolismo , Células Asesinas Naturales/metabolismo , Activación de Linfocitos/inmunología , Masculino , Ratones , Ratones Endogámicos C57BL , Subfamilia K de Receptores Similares a Lectina de Células NK/inmunología , Subfamilia K de Receptores Similares a Lectina de Células NK/metabolismo , Células T Asesinas Naturales/metabolismo , Proteína de la Leucemia Promielocítica con Dedos de Zinc/inmunología , Proteína de la Leucemia Promielocítica con Dedos de Zinc/metabolismo , Receptores de Antígenos de Linfocitos T alfa-beta/inmunología , Receptores de Antígenos de Linfocitos T alfa-beta/metabolismo , Subgrupos de Linfocitos T/inmunología , Subgrupos de Linfocitos T/metabolismoRESUMEN
Innate lymphoid cells (ILC) are a subset of leukocytes with lymphoid properties that lack antigen specific receptors. They can be stimulated by and exert their effect via specific cytokine axes, whereas Natural Killers (NK) cells are the only known cytotoxic member of this family. ILCs are considered key in linking the innate and adaptive response in physiologic and pathologic environments. In this study, we investigated the properties of non-cytotoxic cardiac ILCs in physiologic, inflammatory, and ischemic conditions. We found that in healthy humans and mice, non-cytotoxic cardiac ILCs are predominantly a type 2-committed population with progenitor-like features, such as an absence of type-specific immunophenotype, intermediate GATA3 expression, and capacity to transiently express Pro-myelocytic Leukemia Zinc Finger protein (PLZF) upon activation. During myocarditis and ischemia, in both human and mice, cardiac ILCs differentiated into conventional ILC2s. We found that cardiac ILCs lack IL-25 receptor and cannot become inflammatory ILC2s. We found a strong correlation between IL-33 production in the heart and the ability of cardiac ILCs to become conventional ILC2s. The main producer of IL-33 was a subset of CD29+Sca-1+ cardiac fibroblasts. ILC2 expansion and fibroblast-derived IL-33 production were significantly increased in the heart in mouse models of infarction and myocarditis. Despite its progenitor-like status in healthy hearts, cardiac ILCs were unable to become ILC1 or ILC3 in vivo and in vitro. Using adoptive transfer and parabiosis, we demonstrated that the heart, unlike other organs such as lung, cannot be infiltrated by circulating ILCs in adulthood even during cardiac inflammation or ischemia. Thus, the ILC2s present during inflammatory conditions are derived from the heart-resident and quiescent steady-state population. Non-cytotoxic cardiac ILCs are a resident population of ILC2-commited cells, with undifferentiated progenitor-like features in steady-state conditions and an ability to expand and develop pro-inflammatory type 2 features during inflammation or ischemia.
Asunto(s)
Inmunidad Innata , Células Asesinas Naturales/inmunología , Isquemia Miocárdica/inmunología , Miocarditis/inmunología , Miocardio/inmunología , Animales , Femenino , Fibroblastos/inmunología , Fibroblastos/patología , Factor de Transcripción GATA3/inmunología , Regulación de la Expresión Génica/inmunología , Humanos , Interleucina-33/inmunología , Células Asesinas Naturales/patología , Masculino , Ratones , Ratones Noqueados , Isquemia Miocárdica/patología , Miocarditis/patología , Miocardio/patología , Proteína de la Leucemia Promielocítica con Dedos de Zinc/inmunologíaRESUMEN
Cellular metabolism and signaling pathways are key regulators to determine conventional T cell fate and function, but little is understood about the role of cell metabolism for natural killer T (NKT) cell survival, proliferation, and function. We found that NKT cells operate distinct metabolic programming from CD4 T cells. NKT cells are less efficient in glucose uptake than CD4 T cells with or without activation. Gene-expression data revealed that, in NKT cells, glucose is preferentially metabolized by the pentose phosphate pathway and mitochondria, as opposed to being converted into lactate. In fact, glucose is essential for the effector functions of NKT cells and a high lactate environment is detrimental for NKT cell survival and proliferation. Increased glucose uptake and IFN-γ expression in NKT cells is inversely correlated with bacterial loads in response to bacterial infection, further supporting the significance of glucose metabolism for NKT cell function. We also found that promyelocytic leukemia zinc finger seemed to play a role in regulating NKT cells' glucose metabolism. Overall, our study reveals that NKT cells use distinct arms of glucose metabolism for their survival and function.
Asunto(s)
Linfocitos T CD4-Positivos/inmunología , Proliferación Celular , Mitocondrias/metabolismo , Células T Asesinas Naturales/inmunología , Fosforilación Oxidativa , Vía de Pentosa Fosfato/inmunología , Animales , Linfocitos T CD4-Positivos/citología , Supervivencia Celular/genética , Supervivencia Celular/inmunología , Glucosa/genética , Glucosa/inmunología , Ratones , Ratones Noqueados , Mitocondrias/genética , Células T Asesinas Naturales/citología , Vía de Pentosa Fosfato/genética , Proteína de la Leucemia Promielocítica con Dedos de Zinc/genética , Proteína de la Leucemia Promielocítica con Dedos de Zinc/inmunologíaRESUMEN
Expansion of natural killer (NK) cells expressing NKG2C occurs following human cytomegalovirus (HCMV) infection and is amplified by human immunodeficiency virus (HIV) co-infection. These NKG2C-expressing NK cells demonstrate enhanced CD16-dependent cytokine production and downregulate FcεRIγ and promyelocytic leukemia zinc finger protein (PLZF). Lacking NKG2C diminishes resistance to HIV infection, but whether this affects NK cell acquisition of superior antibody-dependent function is unclear. Therefore, our objective was to investigate whether HCMV-driven NK cell differentiation is impaired in NKG2Cnull HIV-infected individuals. Phenotypic (CD2, CD16, CD57, NKG2A, FcεRIγ, and PLZF expression) and functional (cytokine induction and cytotoxicity) properties were compared between HIVâ»infected NKG2Cnull and NKG2C-expressing groups. Cytokine production was compared following stimulation through natural cytotoxicity receptors or through CD16. Cytotoxicity was measured by anti-CD16-redirected lysis and by classical antibody-dependent cell-mediated cytotoxicity (ADCC) against anti-class I human leukocyte antigen (HLA) antibody-coated cells. Our data indicate highly similar HCMV-driven NK cell differentiation in HIV infection with or without NKG2C. While the fraction of mature (CD57pos) NK cells expressing CD2 (p = 0.009) or co-expressing CD2 and CD16 (p = 0.03) was significantly higher in NKG2Cnull HIV-infected individuals, there were no significant differences in NKG2A, FcεRIγ, or PLZF expression. The general phenotypic and functional equivalency observed suggests NKG2C-independent routes of HCMV-driven NK cell differentiation, which may involve increased CD2 expression.
Asunto(s)
Infecciones por Citomegalovirus/inmunología , Citomegalovirus/patogenicidad , Infecciones por VIH/inmunología , Células Asesinas Naturales/virología , Subfamília C de Receptores Similares a Lectina de Células NK/genética , Adulto , Citotoxicidad Celular Dependiente de Anticuerpos , Diferenciación Celular , Coinfección/inmunología , Coinfección/virología , Citocinas/inmunología , Femenino , Proteínas Ligadas a GPI/genética , Proteínas Ligadas a GPI/inmunología , Humanos , Células Asesinas Naturales/citología , Activación de Linfocitos , Masculino , Persona de Mediana Edad , Subfamília C de Receptores Similares a Lectina de Células NK/inmunología , Proteína de la Leucemia Promielocítica con Dedos de Zinc/genética , Proteína de la Leucemia Promielocítica con Dedos de Zinc/inmunología , Receptores de IgE/genética , Receptores de IgE/inmunología , Receptores de IgG/genética , Receptores de IgG/inmunologíaRESUMEN
Multipotent progenitor cells confirm their T cell-lineage identity in the CD4-CD8- double-negative (DN) pro-T cell DN2 stages, when expression of the essential transcription factor Bcl11b begins. In vivo and in vitro stage-specific deletions globally identified Bcl11b-controlled target genes in pro-T cells. Proteomics analysis revealed that Bcl11b associated with multiple cofactors and that its direct action was needed to recruit those cofactors to selective target sites. Regions near functionally regulated target genes showed enrichment for those sites of Bcl11b-dependent recruitment of cofactors, and deletion of individual cofactors relieved the repression of many genes normally repressed by Bcl11b. Runx1 collaborated with Bcl11b most frequently for both activation and repression. In parallel, Bcl11b indirectly regulated a subset of target genes by a gene network circuit via the transcription inhibitor Id2 (encoded by Id2) and transcription factor PLZF (encoded by Zbtb16); Id2 and Zbtb16 were directly repressed by Bcl11b, and Id2 and PLZF controlled distinct alternative programs. Thus, our study defines the molecular basis of direct and indirect Bcl11b actions that promote T cell identity and block alternative potentials.
Asunto(s)
Linfopoyesis/inmunología , Células Precursoras de Linfocitos T/inmunología , Proteína de la Leucemia Promielocítica con Dedos de Zinc/biosíntesis , Proteínas Represoras/inmunología , Proteínas Supresoras de Tumor/inmunología , Animales , Diferenciación Celular/inmunología , Regulación de la Expresión Génica/inmunología , Proteína 2 Inhibidora de la Diferenciación/biosíntesis , Proteína 2 Inhibidora de la Diferenciación/inmunología , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Células Precursoras de Linfocitos T/citología , Proteína de la Leucemia Promielocítica con Dedos de Zinc/inmunologíaAsunto(s)
Inmunidad Innata/inmunología , Proteína de la Leucemia Promielocítica con Dedos de Zinc/inmunología , Timocitos/inmunología , Animales , Diferenciación Celular/inmunología , Receptores de Hialuranos/inmunología , Interferón gamma/inmunología , Interleucina-4/inmunología , Ratones , Células T Asesinas Naturales/inmunologíaRESUMEN
γδ NKT cells are neonatal-derived γδ T lymphocytes that are grouped together with invariant NKT cells based on their shared innate-like developmental program characterized by the transcription factor PLZF (promyelocytic leukemia zinc finger). Previous studies have demonstrated that the population size of γδ NKT cells is tightly controlled by Id3-mediated inhibition of E-protein activity in mice. However, how E proteins promote γδ NKT cell development and expansion remains to be determined. In this study, we report that the transcription factor Egr2, which also activates PLZF expression in invariant NKT cells, is essential for regulating γδ NKT cell expansion. We observed a higher expression of Egr family genes in γδ NKT cells compared with the conventional γδ T cell population. Loss of function of Id3 caused an expansion of γδ NKT cells, which is accompanied by further upregulation of Egr family genes as well as PLZF. Deletion of Egr2 in Id3-deficient γδ NKT cells prevented cell expansion and blocked PLZF upregulation. We further show that this Egr2-mediated γδ NKT cell expansion is dependent on c-Myc. c-Myc knockdown attenuated the proliferation of Id3-deficient γδ NKT cells, whereas c-Myc overexpression enhanced the proliferation of Id3/Egr2-double-deficient γδ NKT cells. Therefore, our data reveal a regulatory circuit involving Egr2-Id3-E2A, which normally restricts the population size of γδ NKT cells by adjusting Egr2 dosage and c-Myc expression.
Asunto(s)
Proliferación Celular/fisiología , Proteína 2 de la Respuesta de Crecimiento Precoz/inmunología , Regulación de la Expresión Génica/inmunología , Proteínas Inhibidoras de la Diferenciación/inmunología , Células T Asesinas Naturales/inmunología , Proteínas Proto-Oncogénicas c-myc/inmunología , Animales , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/genética , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/inmunología , Proteína 2 de la Respuesta de Crecimiento Precoz/genética , Proteínas Inhibidoras de la Diferenciación/genética , Ratones , Ratones Noqueados , Células T Asesinas Naturales/citología , Proteína de la Leucemia Promielocítica con Dedos de Zinc/genética , Proteína de la Leucemia Promielocítica con Dedos de Zinc/inmunología , Proteínas Proto-Oncogénicas c-myc/genética , Receptores de Antígenos de Linfocitos T gamma-deltaRESUMEN
Zbtb16-encoded PLZF is a signature transcription factor (TF) that directs the acquisition of T-helper effector programs during the development of multiple innate lymphocyte lineages, including natural killer T cell, innate lymphoid cell, mucosal-associated invariant T cell and γδ lineages. PLZF is also essential in osteoblast and spermatogonial development. How Zbtb16 itself is regulated in different lineages is incompletely understood. Here, by systematic CRISPR/Cas9-assisted deletions of chromatin accessible regions within the Zbtb16 locus in mouse, we identify a critical enhancer controlling PLZF expression exclusively in innate lymphoid lineages. Multiple sites within this enhancer express canonical motifs for the TF Runx1, which is essential for the development of these lineages. Notably, some regulatory sites control the kinetic rather than the overall level of PLZF expression. Thus, our comprehensive, unbiased analysis of regulatory elements in vivo reveals critical mechanisms of Zbtb16 regulation shared between innate and innate-like lymphoid lineages. Zbtb16-encoded transcription factor PLZF directs the differentiation of multiple innate and innate-like cell lineages, but how Zbtb16 itself is regulated remains unclear. Here the authors show, using CRISPR gene editing, ATAC-seq and ChIP-seq, that specific Runx1-bound enhancer elements critically modulate lineage-dependent expressions of PLZF.
Asunto(s)
Subunidad alfa 2 del Factor de Unión al Sitio Principal/genética , Linfocitos Intraepiteliales/inmunología , Linfocitos/inmunología , Células T Asesinas Naturales/inmunología , Proteína de la Leucemia Promielocítica con Dedos de Zinc/genética , Linfocitos T Colaboradores-Inductores/inmunología , Animales , Sistemas CRISPR-Cas , Diferenciación Celular , Linaje de la Célula , Elementos de Facilitación Genéticos , Epigénesis Genética , Inmunidad Innata/inmunología , Linfocitos Intraepiteliales/citología , Linfocitos/citología , Linfopoyesis , Ratones , Ratones Noqueados , Células T Asesinas Naturales/citología , Proteína de la Leucemia Promielocítica con Dedos de Zinc/inmunología , Secuencias Reguladoras de Ácidos Nucleicos , Linfocitos T Colaboradores-Inductores/citologíaRESUMEN
The development of CD1d-restricted invariant natural killer T (iNKT) cells, a population that is critical for both innate and adaptive immunity, is regulated by multiple transcription factors, but the molecular mechanisms underlying how the transcriptional activation of these factors are regulated during iNKT development remain largely unknown. We found that the histone acetyltransferase general control non-derepressible 5 (GCN5) is essential for iNKT cell development during the maturation stage. GCN5 deficiency blocked iNKT cell development in a cell-intrinsic manner. At the molecular level, GCN5 is a specific lysine acetyltransferase of early growth responsive gene 2 (EGR2), a transcription factor required for iNKT cell development. GCN5-mediated acetylation positively regulated EGR2 transcriptional activity, and both genetic and pharmacological GCN5 suppression specifically inhibited the transcription of EGR2 target genes in iNKT cells, including Runx1, promyelocytic leukemia zinc finger protein (PLZF), interleukin (IL)-2Rb, and T-bet. Therefore, our study revealed GCN5-mediated EGR2 acetylation as a molecular mechanism that regulates iNKT development.