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1.
Cell ; 164(3): 365-77, 2016 Jan 28.
Artículo en Inglés | MEDLINE | ID: mdl-26806130

RESUMEN

Malignancy can be suppressed by the immune system in a process termed immunosurveillance. However, to what extent immunosurveillance occurs in spontaneous cancers and the composition of participating cell types remains obscure. Here, we show that cell transformation triggers a tissue-resident lymphocyte response in oncogene-induced murine cancer models. Non-circulating cytotoxic lymphocytes, derived from innate, T cell receptor (TCR)αß, and TCRγδ lineages, expand in early tumors. Characterized by high expression of NK1.1, CD49a, and CD103, these cells share a gene-expression signature distinct from those of conventional NK cells, T cells, and invariant NKT cells. Generation of these lymphocytes is dependent on the cytokine IL-15, but not the transcription factor Nfil3 that is required for the differentiation of tumor-infiltrating NK cells, and IL-15 deficiency, but not Nfil3 deficiency, results in accelerated tumor growth. These findings reveal a tumor-elicited immunosurveillance mechanism that engages unconventional type-1-like innate lymphoid cells and type 1 innate-like T cells.


Asunto(s)
Linfocitos/inmunología , Neoplasias Mamarias Experimentales/inmunología , Monitorización Inmunológica , Subgrupos de Linfocitos T/inmunología , Animales , Factores de Transcripción con Cremalleras de Leucina de Carácter Básico/metabolismo , Granzimas/metabolismo , Interleucina-15/inmunología , Ratones , Ratones Endogámicos C57BL , Receptores de Antígenos de Linfocitos T alfa-beta/metabolismo
2.
Immunity ; 43(2): 211-3, 2015 Aug 18.
Artículo en Inglés | MEDLINE | ID: mdl-26287675

RESUMEN

Demethylation of the Foxp3 locus maintains gene expression and Treg cell stability. Yang et al. (2015) show that the gasotransmitter hydrogen sulfide co-operates with growth factor TGF-ß and interleukin-2 to activate Tet-mediated DNA demethylation of Foxp3 to promote immune tolerance.


Asunto(s)
Colitis/inmunología , Proteínas de Unión al ADN/metabolismo , Factores de Transcripción Forkhead/metabolismo , Sulfuro de Hidrógeno/metabolismo , Proteínas Proto-Oncogénicas/metabolismo , Linfocitos T Reguladores/inmunología , Animales , Humanos
3.
Immunity ; 39(2): 335-46, 2013 Aug 22.
Artículo en Inglés | MEDLINE | ID: mdl-23932572

RESUMEN

Interleukin-7 receptor α chain (IL-7Rα) is induced upon T cell positive selection and controls thymic CD8-lineage specification and peripheral naive T cell homeostasis. How IL-7Rα expression is regulated in developing thymocytes is unclear. Here, we show that transforming growth factor ß (TGF-ß) signaling promoted IL-7Rα expression and CD8+ T cell differentiation. In addition, TGF-ß signaling was required for high IL-7Rα expression in CD4+ T cells bearing low-affinity T cell receptors, and the abrogation of TGF-ß receptor expression led to failed maintenance of peripheral CD4+ T cells. Compromised IL-7Rα expression in TGF-ß-receptor-deficient T cells was associated with increased expression of the Il7ra transcriptional repressor, Gfi-1. IL-7Rα transgenesis or T-cell-specific ablation of Gfi-1 restored IL-7Rα expression and largely ameliorated the development and homeostasis defects of TGF-ß-receptor-deficient T cells. These findings reveal functions for TGF-ß signaling in controlling IL-7Rα expression and in promoting T cell repertoire diversification.


Asunto(s)
Linfocitos T CD4-Positivos/inmunología , Linfocitos T CD8-positivos/metabolismo , Proteínas de Unión al ADN/metabolismo , Receptores de Interleucina-7/metabolismo , Factores de Transcripción/metabolismo , Factor de Crecimiento Transformador beta/metabolismo , Animales , Linfocitos T CD8-positivos/inmunología , Diferenciación Celular , Linaje de la Célula , Proteínas de Unión al ADN/genética , Homeostasis , Ratones , Ratones Noqueados , Proteínas Serina-Treonina Quinasas/deficiencia , Proteínas Serina-Treonina Quinasas/genética , Proteínas Serina-Treonina Quinasas/metabolismo , Receptor Tipo II de Factor de Crecimiento Transformador beta , Receptores de Antígenos de Linfocitos T , Receptores de Interleucina-7/inmunología , Receptores de Factores de Crecimiento Transformadores beta/deficiencia , Receptores de Factores de Crecimiento Transformadores beta/genética , Receptores de Factores de Crecimiento Transformadores beta/metabolismo , Transducción de Señal , Factores de Transcripción/genética
4.
Proc Natl Acad Sci U S A ; 114(36): E7536-E7544, 2017 09 05.
Artículo en Inglés | MEDLINE | ID: mdl-28827353

RESUMEN

Peripheral T cell tolerance is promoted by the regulatory cytokine TGF-ß and Foxp3-expressing Treg cells. However, whether TGF-ß and Treg cells are part of the same regulatory module, or exist largely as distinct pathways to repress self-reactive T cells remains incompletely understood. Using a transgenic model of autoimmune diabetes, here we show that ablation of TGF-ß receptor II (TßRII) in T cells, but not Foxp3 deficiency, resulted in early-onset diabetes with complete penetrance. The rampant autoimmune disease was associated with enhanced T cell priming and elevated T cell expression of the inflammatory cytokine GM-CSF, concomitant with pancreatic infiltration of inflammatory monocytes that triggered immunopathology. Ablation of the GM-CSF receptor alleviated the monocyte response and inhibited disease development. These findings reveal that TGF-ß promotes T cell tolerance primarily via Foxp3-independent mechanisms and prevents autoimmunity in this model by repressing the cross talk between adaptive and innate immune systems.


Asunto(s)
Factores de Transcripción Forkhead/metabolismo , Tolerancia Periférica/inmunología , Linfocitos T Reguladores/metabolismo , Factor de Crecimiento Transformador beta/metabolismo , Inmunidad Adaptativa/inmunología , Animales , Autoinmunidad/inmunología , Citocinas/metabolismo , Factor Estimulante de Colonias de Granulocitos y Macrófagos/metabolismo , Inmunidad Innata/inmunología , Inflamación/metabolismo , Ratones , Receptores de Factor Estimulante de Colonias de Granulocitos y Macrófagos/metabolismo , Linfocitos T Reguladores/inmunología
5.
J Immunol ; 191(8): 3973-9, 2013 Oct 15.
Artículo en Inglés | MEDLINE | ID: mdl-24098055

RESUMEN

A fundamental aspect of the adaptive immune system is the generation and maintenance of a diverse and self-tolerant T cell repertoire. Through its regulation of T cell development, homeostasis, tolerance, and differentiation, the highly evolutionarily conserved cytokine TGF-ß critically supports a functional T cell pool. The pleiotropic nature of this regulation is likely due to the elaborate control of TGF-ß production and activation in the immune system, and the intricacy of TGF-ß signaling pathways. In this review we discuss the current understanding of TGF-ß regulation of T cells.


Asunto(s)
Linfocitos T CD4-Positivos/inmunología , Linfocitos T CD8-positivos/inmunología , Factor de Crecimiento Transformador beta/inmunología , Factor de Crecimiento Transformador beta/metabolismo , Animales , Diferenciación Celular , Linaje de la Célula , Activación Enzimática , Humanos , Activación de Linfocitos , Ratones , Autotolerancia , Transducción de Señal
6.
Nat Biotechnol ; 2023 Oct 19.
Artículo en Inglés | MEDLINE | ID: mdl-37857725

RESUMEN

The broad application of precision cancer immunotherapies is limited by the number of validated neoepitopes that are common among patients or tumor types. To expand the known repertoire of shared neoantigen-human leukocyte antigen (HLA) complexes, we developed a high-throughput platform that coupled an in vitro peptide-HLA binding assay with engineered cellular models expressing individual HLA alleles in combination with a concatenated transgene harboring 47 common cancer neoantigens. From more than 24,000 possible neoepitope-HLA combinations, biochemical and computational assessment yielded 844 unique candidates, of which 86 were verified after immunoprecipitation mass spectrometry analyses of engineered, monoallelic cell lines. To evaluate the potential for immunogenicity, we identified T cell receptors that recognized select neoepitope-HLA pairs and elicited a response after introduction into human T cells. These cellular systems and our data on therapeutically relevant neoepitopes in their HLA contexts will aid researchers studying antigen processing as well as neoepitope targeting therapies.

7.
Curr Protoc ; 2(9): e538, 2022 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-36130036

RESUMEN

Effective and precise gene editing of T lymphocytes is critical for advancing the understanding of T cell biology and the development of next-generation cellular therapies. Although methods for effective CRISPR/Cas9-mediated gene knock-out in primary human T cells have been developed, complementary techniques for nonviral gene knock-in can be cumbersome and inefficient. Here, we report a simple and efficient method for nonviral CRISPR/Cas9-based gene knock-in utilizing plasmid-based donor DNA templates. © 2022 Wiley Periodicals LLC. Basic Protocol 1: Purification of human CD4+ or CD8+ T cells from blood Basic Protocol 2: Activation of purified CD4+ or CD8+ T cells using TransAct CD3/CD28 agonist-conjugated nanomatrix Basic Protocol 3: Preparation of Cas9/sgRNA RNPs Basic Protocol 4: Transfection of CAS9-RNP and knock-in template into human T cells Support Protocol 1: Purity check following magnetic T cell isolation Support Protocol 2: Dextramer staining of TCR-edited T cells Support Protocol 3: Functional characterization of TCR knock-in T cells Support Protocol 4: Detection of knock-in reporter activity in CRISPR/CAS9-edited T cells.


Asunto(s)
Linfocitos T CD8-positivos , Sistemas CRISPR-Cas , Antígenos CD28/genética , Linfocitos T CD8-positivos/metabolismo , Sistemas CRISPR-Cas/genética , ADN , Técnicas de Sustitución del Gen , Humanos , Plásmidos/genética , Receptores de Antígenos de Linfocitos T/genética , Ribonucleoproteínas/genética
8.
J Exp Med ; 219(5)2022 05 02.
Artículo en Inglés | MEDLINE | ID: mdl-35452075

RESUMEN

Genome engineering of T lymphocytes, the main effectors of antitumor adaptive immune responses, has the potential to uncover unique insights into their functions and enable the development of next-generation adoptive T cell therapies. Viral gene delivery into T cells, which is currently used to generate CAR T cells, has limitations in regard to targeting precision, cargo flexibility, and reagent production. Nonviral methods for effective CRISPR/Cas9-mediated gene knock-out in primary human T cells have been developed, but complementary techniques for nonviral gene knock-in can be cumbersome and inefficient. Here, we report a convenient and scalable nonviral method that allows precise gene edits and transgene integration in primary human T cells, using plasmid donor DNA template and Cas9-RNP. This method is highly efficient for single and multiplex gene manipulation, without compromising T cell function, and is thus valuable for use in basic and translational research.


Asunto(s)
Sistemas CRISPR-Cas , Edición Génica , Sistemas CRISPR-Cas/genética , ADN/genética , Edición Génica/métodos , Humanos , Plásmidos/genética , Linfocitos T
9.
Nat Cancer ; 1(7): 681-691, 2020 07.
Artículo en Inglés | MEDLINE | ID: mdl-35122038

RESUMEN

Inhibiting the programmed death-1 (PD-1) pathway is one of the most effective approaches to cancer immunotherapy, but its mechanistic basis remains incompletely understood. Binding of PD-1 to its ligand PD-L1 suppresses T-cell function in part by inhibiting CD28 signaling. Tumor cells and infiltrating myeloid cells can express PD-L1, with myeloid cells being of particular interest as they also express B7-1, a ligand for CD28 and PD-L1. Here we demonstrate that dendritic cells (DCs) represent a critical source of PD-L1, despite being vastly outnumbered by PD-L1+ macrophages. Deletion of PD-L1 in DCs, but not macrophages, greatly restricted tumor growth and led to enhanced antitumor CD8+ T-cell responses. Our data identify a unique role for DCs in the PD-L1-PD-1 regulatory axis and have implications for understanding the therapeutic mechanism of checkpoint blockade, which has long been assumed to reflect the reversal of T-cell exhaustion induced by PD-L1+ tumor cells.


Asunto(s)
Antígeno B7-H1 , Neoplasias , Antígeno B7-H1/genética , Antígenos CD28/metabolismo , Células Dendríticas , Humanos , Ligandos , Neoplasias/genética , Receptor de Muerte Celular Programada 1/genética
10.
Curr Protoc Immunol ; 124(1): e69, 2019 02.
Artículo en Inglés | MEDLINE | ID: mdl-30334617

RESUMEN

CRISPR/Cas9 has enabled the rapid and efficient generation of gene knockouts across various cell types of several species. T cells are central players in adaptive immune responses. Gene editing in primary T cells not only represents a valuable research tool, but is also critical for next generation immunotherapies, such as CAR T cells. Broad application of CRIPSR/Cas9 for gene editing in primary T cells has been hampered by limitations in transfection efficiency and the requirement for TCR stimulation. In this article, we provide a detailed protocol for Cas9/gRNA ribonucleoprotein (RNP) transfection of primary mouse and human T cells without the need for TCR stimulation that achieves near complete loss of target gene expression at the population level. This approach enables rapid target discovery and validation in both mouse and human primary T cells. © 2018 by John Wiley & Sons, Inc.


Asunto(s)
Linfocitos T CD4-Positivos/metabolismo , Linfocitos T CD8-positivos/metabolismo , Sistemas CRISPR-Cas/genética , Ribonucleoproteínas/genética , Animales , Linfocitos T CD4-Positivos/citología , Linfocitos T CD8-positivos/citología , Edición Génica , Humanos , Ratones , Ratones Noqueados , Ribonucleoproteínas/metabolismo , Transfección
11.
Artículo en Inglés | MEDLINE | ID: mdl-28108486

RESUMEN

Transforming growth factor ß (TGF-ß) is a pleiotropic cytokine involved in both suppressive and inflammatory immune responses. After 30 years of intense study, we have only begun to elucidate how TGF-ß alters immunity under various conditions. Under steady-state conditions, TGF-ß regulates thymic T-cell selection and maintains homeostasis of the naïve T-cell pool. TGF-ß inhibits cytotoxic T lymphocyte (CTL), Th1-, and Th2-cell differentiation while promoting peripheral (p)Treg-, Th17-, Th9-, and Tfh-cell generation, and T-cell tissue residence in response to immune challenges. Similarly, TGF-ß controls the proliferation, survival, activation, and differentiation of B cells, as well as the development and functions of innate cells, including natural killer (NK) cells, macrophages, dendritic cells, and granulocytes. Collectively, TGF-ß plays a pivotal role in maintaining peripheral tolerance against self- and innocuous antigens, such as food, commensal bacteria, and fetal alloantigens, and in controlling immune responses to pathogens.


Asunto(s)
Autoinmunidad , Activación de Linfocitos , Linfocitos T Reguladores/inmunología , Factor de Crecimiento Transformador beta/inmunología , Animales , Artritis Reumatoide/inmunología , Linfocitos B/inmunología , Infecciones Bacterianas/inmunología , Diferenciación Celular , Linaje de la Célula , Proliferación Celular , Supervivencia Celular , Células Dendríticas/inmunología , Diabetes Mellitus Tipo 1/inmunología , Granulocitos/inmunología , Homeostasis , Humanos , Tolerancia Inmunológica , Enfermedades Inflamatorias del Intestino/inmunología , Isoantígenos/inmunología , Células Asesinas Naturales/inmunología , Lupus Eritematoso Sistémico/inmunología , Macrófagos/inmunología , Mastocitos/inmunología , Ratones , Monocitos/inmunología , Enfermedades Parasitarias/inmunología , Linfocitos T/inmunología , Timo/inmunología
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