RESUMEN
Respiratory infections cause significant morbidity and mortality, yet it is unclear why some individuals succumb to severe disease. In patients hospitalized with avian A(H7N9) influenza, we investigated early drivers underpinning fatal disease. Transcriptomics strongly linked oleoyl-acyl-carrier-protein (ACP) hydrolase (OLAH), an enzyme mediating fatty acid production, with fatal A(H7N9) early after hospital admission, persisting until death. Recovered patients had low OLAH expression throughout hospitalization. High OLAH levels were also detected in patients hospitalized with life-threatening seasonal influenza, COVID-19, respiratory syncytial virus (RSV), and multisystem inflammatory syndrome in children (MIS-C) but not during mild disease. In olah-/- mice, lethal influenza infection led to survival and mild disease as well as reduced lung viral loads, tissue damage, infection-driven pulmonary cell infiltration, and inflammation. This was underpinned by differential lipid droplet dynamics as well as reduced viral replication and virus-induced inflammation in macrophages. Supplementation of oleic acid, the main product of OLAH, increased influenza replication in macrophages and their inflammatory potential. Our findings define how the expression of OLAH drives life-threatening viral disease.
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COVID-19 , Gripe Humana , Animales , Humanos , Ratones , COVID-19/virología , COVID-19/genética , Gripe Humana/virología , Replicación Viral , Macrófagos/metabolismo , Macrófagos/virología , Femenino , Masculino , SARS-CoV-2 , Pulmón/virología , Pulmón/patología , Pulmón/metabolismo , Ratones Endogámicos C57BL , Ácido Oléico/metabolismo , Infecciones por Virus Sincitial Respiratorio/virología , Ratones Noqueados , Carga Viral , Hidrolasas de Éster Carboxílico/metabolismo , Hidrolasas de Éster Carboxílico/genética , Infecciones por Orthomyxoviridae/virología , Infecciones del Sistema Respiratorio/virología , NiñoRESUMEN
Sepsis is a biphasic disease characterized by an acute inflammatory response, followed by a prolonged immunosuppressive phase. Therapies aimed at controlling inflammation help to reduce the time patients with sepsis spend in intensive care units, but they do not lead to a reduction in overall mortality. Recently, the focus has been on addressing the immunosuppressive phase, often caused by apoptosis of immune cells. However, molecular triggers of these events are not yet known. Using whole-genome CRISPR screening in mice, we identified a triggering receptor expressed on myeloid cells (TREM) family receptor, TREML4, as a key regulator of inflammation and immune cell death in sepsis. Genetic ablation of Treml4 in mice demonstrated that TREML4 regulates calcium homeostasis, the inflammatory cytokine response, myeloperoxidase activation, the endoplasmic reticulum stress response and apoptotic cell death in innate immune cells, leading to an overall increase in survival rate, both during the acute and chronic phases of polymicrobial sepsis.
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Susceptibilidad a Enfermedades , Inmunidad Innata , Receptores Inmunológicos/genética , Receptores Inmunológicos/metabolismo , Sepsis/etiología , Animales , Biomarcadores , Muerte Celular , Repeticiones Palindrómicas Cortas Agrupadas y Regularmente Espaciadas , Citocinas/metabolismo , Modelos Animales de Enfermedad , Susceptibilidad a Enfermedades/inmunología , Edición Génica , Técnicas de Silenciamiento del Gen , Marcación de Gen , Genómica/métodos , Inmunofenotipificación , Inflamación/etiología , Inflamación/metabolismo , Ratones , Ratones Noqueados , Neutrófilos/inmunología , Neutrófilos/metabolismo , Fenotipo , Linfocitos T/inmunología , Linfocitos T/metabolismoRESUMEN
Immune-modulating therapies have revolutionized the treatment of chronic diseases, particularly cancer. However, their success is restricted and there is a need to identify new therapeutic targets. Here, we show that natural killer cell granule protein 7 (NKG7) is a regulator of lymphocyte granule exocytosis and downstream inflammation in a broad range of diseases. NKG7 expressed by CD4+ and CD8+ T cells played key roles in promoting inflammation during visceral leishmaniasis and malaria-two important parasitic diseases. Additionally, NKG7 expressed by natural killer cells was critical for controlling cancer initiation, growth and metastasis. NKG7 function in natural killer and CD8+ T cells was linked with their ability to regulate the translocation of CD107a to the cell surface and kill cellular targets, while NKG7 also had a major impact on CD4+ T cell activation following infection. Thus, we report a novel therapeutic target expressed on a range of immune cells with functions in different immune responses.
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Linfocitos T CD4-Positivos/inmunología , Linfocitos T CD8-positivos/inmunología , Inflamación/inmunología , Células Asesinas Naturales/inmunología , Leishmania donovani/fisiología , Leishmaniasis Visceral/inmunología , Malaria/inmunología , Proteínas de la Membrana/metabolismo , Plasmodium/fisiología , Animales , Células Cultivadas , Citotoxicidad Inmunológica , Modelos Animales de Enfermedad , Exocitosis , Humanos , Proteínas de la Membrana/genética , Ratones , Ratones Endogámicos C57BL , Vesículas Secretoras/metabolismoRESUMEN
Programmed cell death contributes to host defense against pathogens. To investigate the relative importance of pyroptosis, necroptosis, and apoptosis during Salmonella infection, we infected mice and macrophages deficient for diverse combinations of caspases-1, -11, -12, and -8 and receptor interacting serine/threonine kinase 3 (RIPK3). Loss of pyroptosis, caspase-8-driven apoptosis, or necroptosis had minor impact on Salmonella control. However, combined deficiency of these cell death pathways caused loss of bacterial control in mice and their macrophages, demonstrating that host defense can employ varying components of several cell death pathways to limit intracellular infections. This flexible use of distinct cell death pathways involved extensive cross-talk between initiators and effectors of pyroptosis and apoptosis, where initiator caspases-1 and -8 also functioned as executioners when all known effectors of cell death were absent. These findings uncover a highly coordinated and flexible cell death system with in-built fail-safe processes that protect the host from intracellular infections.
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Apoptosis/inmunología , Macrófagos/inmunología , Necroptosis/inmunología , Piroptosis/inmunología , Infecciones por Salmonella/inmunología , Salmonella/inmunología , Animales , Caspasa 1/deficiencia , Caspasa 1/genética , Caspasa 12/deficiencia , Caspasa 12/genética , Caspasa 8/genética , Caspasas Iniciadoras/deficiencia , Caspasas Iniciadoras/genética , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Proteína Serina-Treonina Quinasas de Interacción con Receptores/deficiencia , Proteína Serina-Treonina Quinasas de Interacción con Receptores/genéticaRESUMEN
Investigating how transcription factors control complex cellular processes requires tools that enable responses to be visualised at the single-cell level and their cell fate to be followed over time. For example, the tumour suppressor p53 (also called TP53 in humans and TRP53 in mice) can initiate diverse cellular responses by transcriptional activation of its target genes: Puma to induce apoptotic cell death and p21 to induce cell cycle arrest/cell senescence. However, it is not known how these processes are regulated and initiated in different cell types. Also, the context-dependent interaction partners and binding loci of p53 remain largely elusive. To be able to examine these questions, we here developed knock-in mice expressing triple-FLAG-tagged p53 to facilitate p53 pull-down and two p53 response reporter mice, knocking tdTomato and GFP into the Puma/Bbc3 and p21 gene loci, respectively. By crossing these reporter mice into a p53-deficient background, we show that the new reporters reliably inform on p53-dependent and p53-independent initiation of both apoptotic or cell cycle arrest/senescence programs, respectively, in vitro and in vivo.
RESUMEN
Dendritic cells (DCs) are can be broadly divided into conventional (cDC) and plasmacytoid (pDC) subsets. Despite the importance of this lineage diversity, its genetic basis is not fully understood. We found that conditional ablation of the Ets-family transcription factor PU.1 in DC-restricted progenitors led to increased pDC production at the expense of cDCs. PU.1 controlled many of the cardinal functions of DCs, such as antigen presentation by cDCs and type I interferon production by pDCs. Conditional ablation of PU.1 de-repressed the pDC transcriptional signature in cDCs. The combination of genome-wide mapping of PU.1 binding and gene expression analysis revealed a key role for PU.1 in maintaining cDC identity through the induction of the transcriptional regulator DC-SCRIPT. PU.1 activated DC-SCRIPT expression, which in turn promoted cDC formation, particularly of cDC1s, and repressed pDC development. Thus, cDC identity is regulated by a transcriptional node requiring PU.1 and DC-SCRIPT.
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Proteínas de Unión al ADN/metabolismo , Células Dendríticas/fisiología , Proteínas Nucleares/metabolismo , Proteínas Proto-Oncogénicas/metabolismo , Transactivadores/metabolismo , Factores de Transcripción/metabolismo , Animales , Presentación de Antígeno , Diferenciación Celular , Linaje de la Célula , Proteínas de Unión al ADN/genética , Regulación de la Expresión Génica , Células HEK293 , Humanos , Interferón Tipo I/metabolismo , Ratones , Ratones Transgénicos , Proteínas Nucleares/genética , Proteínas Proto-Oncogénicas/genética , Transducción de Señal , Transactivadores/genética , Factores de Transcripción/genética , TranscriptomaRESUMEN
Polymorphisms in NFKB1 that diminish its expression have been linked to human inflammatory diseases and increased risk for epithelial cancers. The underlying mechanisms are unknown, and the link is perplexing given that NF-κB signaling reportedly typically exerts pro-tumorigenic activity. Here we have shown that NF-κB1 deficiency, even loss of a single allele, resulted in spontaneous invasive gastric cancer (GC) in mice that mirrored the histopathological progression of human intestinal-type gastric adenocarcinoma. Bone marrow chimeras revealed that NF-κB1 exerted tumor suppressive functions in both epithelial and hematopoietic cells. RNA-seq analysis showed that NF-κB1 deficiency resulted in aberrant JAK-STAT signaling, which dysregulated expression of effectors of inflammation, antigen presentation, and immune checkpoints. Concomitant loss of STAT1 prevented these immune abnormalities and GC development. These findings provide mechanistic insight into how polymorphisms that attenuate NFKB1 expression predispose humans to epithelial cancers, highlighting the pro-tumorigenic activity of STAT1 and identifying targetable vulnerabilities in GC.
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Regulación Neoplásica de la Expresión Génica , Inflamación/genética , Inflamación/metabolismo , FN-kappa B/deficiencia , Factor de Transcripción STAT1/metabolismo , Neoplasias Gástricas/genética , Neoplasias Gástricas/metabolismo , Animales , Presentación de Antígeno/inmunología , Células de la Médula Ósea/metabolismo , Células de la Médula Ósea/patología , Transformación Celular Neoplásica/genética , Transformación Celular Neoplásica/metabolismo , Modelos Animales de Enfermedad , Células Epiteliales/metabolismo , Células Epiteliales/patología , Redes Reguladoras de Genes , Humanos , Inflamación/patología , Ratones , Ratones Noqueados , Factor de Transcripción STAT1/deficiencia , Neoplasias Gástricas/inmunología , Neoplasias Gástricas/patologíaRESUMEN
RIPK1 is a key regulator of innate immune signalling pathways. To ensure an optimal inflammatory response, RIPK1 is regulated post-translationally by well-characterized ubiquitylation and phosphorylation events, as well as by caspase-8-mediated cleavage1-7. The physiological relevance of this cleavage event remains unclear, although it is thought to inhibit activation of RIPK3 and necroptosis8. Here we show that the heterozygous missense mutations D324N, D324H and D324Y prevent caspase cleavage of RIPK1 in humans and result in an early-onset periodic fever syndrome and severe intermittent lymphadenopathy-a condition we term 'cleavage-resistant RIPK1-induced autoinflammatory syndrome'. To define the mechanism for this disease, we generated a cleavage-resistant Ripk1D325A mutant mouse strain. Whereas Ripk1-/- mice died postnatally from systemic inflammation, Ripk1D325A/D325A mice died during embryogenesis. Embryonic lethality was completely prevented by the combined loss of Casp8 and Ripk3, but not by loss of Ripk3 or Mlkl alone. Loss of RIPK1 kinase activity also prevented Ripk1D325A/D325A embryonic lethality, although the mice died before weaning from multi-organ inflammation in a RIPK3-dependent manner. Consistently, Ripk1D325A/D325A and Ripk1D325A/+ cells were hypersensitive to RIPK3-dependent TNF-induced apoptosis and necroptosis. Heterozygous Ripk1D325A/+ mice were viable and grossly normal, but were hyper-responsive to inflammatory stimuli in vivo. Our results demonstrate the importance of caspase-mediated RIPK1 cleavage during embryonic development and show that caspase cleavage of RIPK1 not only inhibits necroptosis but also maintains inflammatory homeostasis throughout life.
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Caspasa 8/metabolismo , Enfermedades Autoinflamatorias Hereditarias/metabolismo , Mutación , Proteína Serina-Treonina Quinasas de Interacción con Receptores/metabolismo , Animales , Caspasa 3/metabolismo , Femenino , Enfermedades Autoinflamatorias Hereditarias/genética , Enfermedades Autoinflamatorias Hereditarias/patología , Humanos , Quinasas Quinasa Quinasa PAM/genética , Quinasas Quinasa Quinasa PAM/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Linaje , Proteína Serina-Treonina Quinasas de Interacción con Receptores/deficiencia , Proteína Serina-Treonina Quinasas de Interacción con Receptores/genéticaRESUMEN
Mutations in Trp53, prevalent in human cancer, are reported to drive tumorigenesis through dominant-negative effects (DNEs) over wild-type TRP53 function as well as neomorphic gain-of-function (GOF) activity. We show that five TRP53 mutants do not accelerate lymphomagenesis on a TRP53-deficient background but strongly synergize with c-MYC overexpression in a manner that distinguishes the hot spot Trp53 mutations. RNA sequencing revealed that the mutant TRP53 DNE does not globally repress wild-type TRP53 function but disproportionately impacts a subset of wild-type TRP53 target genes. Accordingly, TRP53 mutant proteins impair pathways for DNA repair, proliferation, and metabolism in premalignant cells. This reveals that, in our studies of lymphomagenesis, mutant TRP53 drives tumorigenesis primarily through the DNE, which modulates wild-type TRP53 function in a manner advantageous for neoplastic transformation.
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Carcinogénesis/genética , Mutación , Proteína p53 Supresora de Tumor/genética , Animales , Linfoma/genética , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Proteína p53 Supresora de Tumor/metabolismoRESUMEN
The histone acetyltransferase HBO1 (MYST2, KAT7) is indispensable for postgastrulation development, histone H3 lysine 14 acetylation (H3K14Ac), and the expression of embryonic patterning genes. In this study, we report the role of HBO1 in regulating hematopoietic stem cell function in adult hematopoiesis. We used 2 complementary cre-recombinase transgenes to conditionally delete Hbo1 (Mx1-Cre and Rosa26-CreERT2). Hbo1-null mice became moribund due to hematopoietic failure with pancytopenia in the blood and bone marrow 2 to 6 weeks after Hbo1 deletion. Hbo1-deleted bone marrow cells failed to repopulate hemoablated recipients in competitive transplantation experiments. Hbo1 deletion caused a rapid loss of hematopoietic progenitors. The numbers of lineage-restricted progenitors for the erythroid, myeloid, B-, and T-cell lineages were reduced. Loss of HBO1 resulted in an abnormally high rate of recruitment of quiescent hematopoietic stem cells (HSCs) into the cell cycle. Cycling HSCs produced progenitors at the expense of self-renewal, which led to the exhaustion of the HSC pool. Mechanistically, genes important for HSC functions were downregulated in HSC-enriched cell populations after Hbo1 deletion, including genes essential for HSC quiescence and self-renewal, such as Mpl, Tek(Tie-2), Gfi1b, Egr1, Tal1(Scl), Gata2, Erg, Pbx1, Meis1, and Hox9, as well as genes important for multipotent progenitor cells and lineage-specific progenitor cells, such as Gata1. HBO1 was required for H3K14Ac through the genome and particularly at gene loci required for HSC quiescence and self-renewal. Our data indicate that HBO1 promotes the expression of a transcription factor network essential for HSC maintenance and self-renewal in adult hematopoiesis.
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Autorrenovación de las Células , Células Madre Hematopoyéticas , Histona Acetiltransferasas , Animales , Células Cultivadas , Senescencia Celular , Eliminación de Gen , Hematopoyesis , Células Madre Hematopoyéticas/citología , Células Madre Hematopoyéticas/metabolismo , Histona Acetiltransferasas/genética , Histona Acetiltransferasas/metabolismo , Ratones Endogámicos C57BLRESUMEN
AIM: Acyl-coenzyme A dehydrogenase family member 10 (ACAD10) is a mitochondrial protein purported to be involved in the fatty acid oxidation pathway. Metformin is the most prescribed therapy for type 2 diabetes; however, its precise mechanisms of action(s) are still being uncovered. Upregulation of ACAD10 is a requirement for metformin's ability to inhibit growth in cancer cells and extend lifespan in Caenorhabditis elegans. However, it is unknown whether ACAD10 plays a role in metformin's metabolic actions. MATERIALS AND METHODS: We assessed the role for ACAD10 on whole-body metabolism and metformin action by generating ACAD10KO mice on a C57BL/6J background via CRISPR-Cas9 technology. In-depth metabolic phenotyping was conducted in both sexes on a normal chow and high fat-high sucrose diet. RESULTS: Compared with wildtype mice, we detected no difference in body composition, energy expenditure or glucose tolerance in male or female ACAD10KO mice, on a chow diet or high-fat, high-sucrose diet (p ≥ .05). Hepatic mitochondrial function and insulin signalling was not different between genotypes under basal or insulin-stimulated conditions (p ≥ .05). Glucose excursions following acute administration of metformin before a glucose tolerance test were not different between genotypes nor was body composition or energy expenditure altered after 4 weeks of daily metformin treatment (p ≥ .05). Despite the lack of a metabolic phenotype, liver lipidomic analysis suggests ACAD10 depletion influences the abundance of specific ceramide species containing very long chain fatty acids, while metformin treatment altered clusters of cholesterol ester, plasmalogen, phosphatidylcholine and ceramide species. CONCLUSIONS: Loss of ACAD10 does not alter whole-body metabolism or impact the acute or chronic metabolic actions of metformin in this model.
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Diabetes Mellitus Tipo 2 , Metformina , Masculino , Femenino , Ratones , Animales , Diabetes Mellitus Tipo 2/metabolismo , Ratones Endogámicos C57BL , Metformina/farmacología , Glucosa/metabolismo , Insulina , Ceramidas , Sacarosa , Dieta Alta en Grasa/efectos adversosRESUMEN
Stuttering is a common speech disorder that interrupts speech fluency and tends to cluster in families. Typically, stuttering is characterized by speech sounds, words or syllables which may be repeated or prolonged and speech that may be further interrupted by hesitations or 'blocks'. Rare variants in a small number of genes encoding lysosomal pathway proteins have been linked to stuttering. We studied a large four-generation family in which persistent stuttering was inherited in an autosomal dominant manner with disruption of the cortico-basal-ganglia-thalamo-cortical network found on imaging. Exome sequencing of three affected family members revealed the PPID c.808C>T (p.Pro270Ser) variant that segregated with stuttering in the family. We generated a Ppid p.Pro270Ser knock-in mouse model and performed ex vivo imaging to assess for brain changes. Diffusion-weighted MRI in the mouse revealed significant microstructural changes in the left corticospinal tract, as previously implicated in stuttering. Quantitative susceptibility mapping also detected changes in cortico-striatal-thalamo-cortical loop tissue composition, consistent with findings in affected family members. This is the first report to implicate a chaperone protein in the pathogenesis of stuttering. The humanized Ppid murine model recapitulates network findings observed in affected family members.
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Tartamudeo , Humanos , Animales , Ratones , Tartamudeo/genética , Tartamudeo/patología , Peptidil-Prolil Isomerasa F , Habla , Encéfalo/diagnóstico por imagen , Encéfalo/patología , Mapeo EncefálicoRESUMEN
The transcription factor Myc is critically important in driving cell proliferation, a function that is frequently dysregulated in cancer. To avoid this dysregulation Myc is tightly controlled by numerous layers of regulation. One such layer is the use of distal regulatory enhancers to drive Myc expression. Here, using chromosome conformation capture to examine B cells of the immune system in the first hours after their activation, we reveal a previously unidentified enhancer of Myc. The interactivity of this enhancer coincides with a dramatic, but discrete, spike in Myc expression 3 h post-activation. However, genetic deletion of this region, has little impact on Myc expression, Myc protein level or in vitro and in vivo cell proliferation. Examination of the enhancer deleted regulatory landscape suggests that enhancer redundancy likely sustains Myc expression. This work highlights not only the importance of temporally examining enhancers, but also the complexity and dynamics of the regulation of critical genes such as Myc.
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Elementos de Facilitación Genéticos , Genes myc , Elementos de Facilitación Genéticos/genética , Factores de Transcripción/metabolismo , Regulación de la Expresión Génica , Regiones Promotoras GenéticasRESUMEN
46,XY gonadal dysgenesis (GD) is a Disorder/Difference of Sex Development (DSD) that can present with phenotypes ranging from ambiguous genitalia to complete male-to-female sex reversal. Around 50% of 46,XY DSD cases receive a molecular diagnosis. In mice, Fibroblast growth factor 9 (FGF9) is an important component of the male sex-determining pathway. Two FGF9 variants reported to date disrupt testis development in mice, but not in humans. Here, we describe a female patient with 46,XY GD harbouring the rare FGF9 variant (missense mutation), NM_002010.2:c.583G > A;p.(Asp195Asn) (D195N). By biochemical and cell-based approaches, the D195N variant disrupts FGF9 protein homodimerisation and FGF9-heparin-binding, and reduces both Sertoli cell proliferation and Wnt4 repression. XY Fgf9D195N/D195N foetal mice show a transient disruption of testicular cord development, while XY Fgf9D195N/- foetal mice show partial male-to-female gonadal sex reversal. In the general population, the D195N variant occurs at an allele frequency of 2.4 × 10-5 , suggesting an oligogenic basis for the patient's DSD. Exome analysis of the patient reveals several known and novel variants in genes expressed in human foetal Sertoli cells at the time of sex determination. Taken together, our results indicate that disruption of FGF9 homodimerization impairs testis determination in mice and, potentially, also in humans in combination with other variants.
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Factor 9 de Crecimiento de Fibroblastos , Disgenesia Gonadal 46 XY , Humanos , Masculino , Femenino , Ratones , Animales , Dimerización , Factor 9 de Crecimiento de Fibroblastos/genética , Testículo , Gónadas , Disgenesia Gonadal 46 XY/genéticaRESUMEN
Suppressor Of Cytokine Signaling (SOCS) 1 is a critical negative regulator of cytokine signaling and required to protect against an excessive inflammatory response. Genetic deletion of Socs1 results in unrestrained cytokine signaling and neonatal lethality, characterised by an inflammatory immune infiltrate in multiple organs. Overexpression and structural studies have suggested that the SOCS1 kinase inhibitory region (KIR) and Src homology 2 (SH2) domain are important for interaction with and inhibition of the receptor-associated JAK1, JAK2 and TYK2 tyrosine kinases, which initiate downstream signaling. To investigate the role of the KIR and SH2 domain in SOCS1 function, we independently mutated key conserved residues in each domain and analysed the impact on cytokine signaling, and the in vivo impact on SOCS1 function. Mutation of the SOCS1-KIR or SH2 domain had no impact on the integrity of the SOCS box complex, however, mutation within the phosphotyrosine binding pocket of the SOCS1-SH2 domain specifically disrupted SOCS1 interaction with phosphorylated JAK1. In contrast, mutation of the KIR did not affect the interaction with JAK1, but did prevent SOCS1 inhibition of JAK1 autophosphorylation. In human and mouse cell lines, both mutants impacted the ability of SOCS1 to restrain cytokine signaling, and crucially, Socs1-R105A and Socs1-F59A mice displayed a neonatal lethality and excessive inflammatory phenotype similar to Socs1-null mice. This study defines a critical and non-redundant role for both the KIR and SH2 domain in endogenous SOCS1 function.
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Citocinas , Proteína 1 Supresora de la Señalización de Citocinas , Dominios Homologos src , Animales , Humanos , Ratones , Citocinas/metabolismo , Fosforilación , Transducción de Señal/fisiología , Proteína 1 Supresora de la Señalización de Citocinas/metabolismo , Proteínas Supresoras de la Señalización de Citocinas/metabolismo , TYK2 Quinasa/metabolismoRESUMEN
The role of RNA-binding proteins of the CCCH-containing family in regulating proinflammatory cytokine production and inflammation is increasingly recognized. We have identified ZC3H12C (Regnase-3) as a potential post-transcriptional regulator of tumor necrosis factor expression and have investigated its role in vivo by generating Zc3h12c-deficient mice that express green fluorescent protein instead of ZC3H12C. Zc3h12c-deficient mice develop hypertrophic lymph nodes. In the immune system, ZC3H12C expression is mostly restricted to the dendritic cell (DC) populations, and we show that DC-restricted ZC3H12C depletion is sufficient to cause lymphadenopathy. ZC3H12C can regulate Tnf messenger RNA stability via its RNase activity in vitro, and we confirmed the role of Tnf in the development of lymphadenopathy. Finally, we found that loss of ZC3H12C did not impact the outcome of skin inflammation in the imiquimod-induced murine model of psoriasis, despite Zc3h12c being identified as a risk factor for psoriasis susceptibility in several genome-wide association studies. Our data suggest a role for ZC3H12C in DC-driven skin homeostasis.
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Linfadenopatía , Psoriasis , Animales , Células Dendríticas , Estudio de Asociación del Genoma Completo , Inflamación/patología , Ganglios Linfáticos/patología , Linfadenopatía/patología , Ratones , Ratones Endogámicos C57BL , Piel/patologíaRESUMEN
The chemokine receptor CXCR3 is expressed on immune cells to co-ordinate lymphocyte activation and migration. CXCR3 binds three chemokine ligands, CXCL9, CXCL10 and CXCL11. These ligands display distinct expression patterns and ligand signaling biases; however, how each ligand functions individually and collaboratively is incompletely understood. CXCL9 and CXCL10 are considered pro-inflammatory chemokines during viral infection, while CXCL11 may induce a tolerizing state. The investigation of the individual role of CXCL11 in vivo has been hampered as C57BL/6 mice carry several mutations that result in a null allele. Here, CRISPR/Cas9 was used to correct these mutations on a C57BL/6 background. It was validated that CXCL11KI mice expressed CXCL11 protein in dendritic cells, spleen and lung. CXCL11KI mice were largely phenotypically indistinguishable from C57BL/6 mice, both at steady-state and during two models of viral infection. While CXCL11 expression did not modify acute antiviral responses, this study provides a new tool to understand the role of CXCL11 in other experimental settings.
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Quimiocina CXCL10 , Quimiocina CXCL11/metabolismo , Virosis , Animales , Quimiocina CXCL10/genética , Quimiocina CXCL11/genética , Quimiocina CXCL9/genética , Quimiocina CXCL9/metabolismo , Inmunidad , Ligandos , Ratones , Ratones Endogámicos C57BLRESUMEN
BCL-2 family proteins regulate the mitochondrial apoptotic pathway. BOK, a multidomain BCL-2 family protein, is generally believed to be an adaptor protein similar to BAK and BAX, regulating the mitochondrial permeability transition during apoptosis. Here we report that BOK is a positive regulator of a key enzyme involved in uridine biosynthesis; namely, uridine monophosphate synthetase (UMPS). Our data suggest that BOK expression enhances UMPS activity, cell proliferation, and chemosensitivity. Genetic deletion of Bok results in chemoresistance to 5-fluorouracil (5-FU) in different cell lines and in mice. Conversely, cancer cells and primary tissues that acquire resistance to 5-FU down-regulate BOK expression. Furthermore, we also provide evidence for a role for BOK in nucleotide metabolism and cell cycle regulation. Our results have implications in developing BOK as a biomarker for 5-FU resistance and have the potential for the development of BOK-mimetics for sensitizing 5-FU-resistant cancers.
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Proteínas Proto-Oncogénicas c-bcl-2/metabolismo , Uridina/metabolismo , Animales , Apoptosis/efectos de los fármacos , Biomarcadores de Tumor/metabolismo , Proliferación Celular/efectos de los fármacos , Daño del ADN , Resistencia a Antineoplásicos/efectos de los fármacos , Fluorouracilo/farmacología , Mamíferos , Ratones , Complejos Multienzimáticos/metabolismo , Orotato Fosforribosiltransferasa/metabolismo , Orotidina-5'-Fosfato Descarboxilasa/metabolismo , Unión Proteica/efectos de los fármacos , Dominios Proteicos , Proteínas Proto-Oncogénicas c-bcl-2/química , Proteína p53 Supresora de Tumor/metabolismoRESUMEN
Somatically acquired mutations in PHF6 (plant homeodomain finger 6) frequently occur in hematopoietic malignancies and often coincide with ectopic expression of TLX3. However, there is no functional evidence to demonstrate whether these mutations contribute to tumorigenesis. Similarly, the role of PHF6 in hematopoiesis is unknown. We report here that Phf6 deletion in mice resulted in a reduced number of hematopoietic stem cells (HSCs), an increased number of hematopoietic progenitor cells, and an increased proportion of cycling stem and progenitor cells. Loss of PHF6 caused increased and sustained hematopoietic reconstitution in serial transplantation experiments. Interferon-stimulated gene expression was upregulated in the absence of PHF6 in hematopoietic stem and progenitor cells. The numbers of hematopoietic progenitor cells and cycling hematopoietic stem and progenitor cells were restored to normal by combined loss of PHF6 and the interferon α and ß receptor subunit 1. Ectopic expression of TLX3 alone caused partially penetrant leukemia. TLX3 expression and loss of PHF6 combined caused fully penetrant early-onset leukemia. Our data suggest that PHF6 is a hematopoietic tumor suppressor and is important for fine-tuning hematopoietic stem and progenitor cell homeostasis.