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1.
Immunity ; 53(4): 775-792.e9, 2020 10 13.
Artigo em Inglês | MEDLINE | ID: mdl-33002412

RESUMO

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.


Assuntos
Imunidade Inata/imunologia , Linfócitos/imunologia , Células Progenitoras Linfoides/imunologia , Animais , Diferenciação Celular/imunologia , Células Cultivadas , Feminino , Humanos , Subunidade alfa de Receptor de Interleucina-18/imunologia , Pulmão/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Proteína com Dedos de Zinco da Leucemia Promielocítica/imunologia , Transdução de Sinais/imunologia , Análise de Célula Única/métodos , Fator 1 de Transcrição de Linfócitos T/imunologia , Fatores de Transcrição/imunologia
2.
Genes Dev ; 34(21-22): 1503-1519, 2020 11 01.
Artigo em Inglês | MEDLINE | ID: mdl-33004416

RESUMO

EBF1 and PAX5 mutations are associated with the development of B progenitor acute lymphoblastic leukemia (B-ALL) in humans. To understand the molecular networks driving leukemia in the Ebf1+/-Pax5+/- (dHet) mouse model for B-ALL, we interrogated the transcriptional profiles and chromatin status of leukemic cells, preleukemic dHet pro-B, and wild-type pro-B cells with the corresponding EBF1 and Pax5 cistromes. In dHet B-ALL cells, many EBF1 and Pax5 target genes encoding pre-BCR signaling components and transcription factors were down-regulated, whereas Myc and genes downstream from IL-7 signaling or associated with the folate pathway were up-regulated. We show that blockade of IL-7 signaling in vivo and methotrexate treatment of leukemic cells in vitro attenuate the expansion of leukemic cells. Single-cell RNA-sequencing revealed heterogeneity of leukemic cells and identified a subset of wild-type pro-B cells with reduced Ebf1 and enhanced Myc expression that show hallmarks of dHet B-ALL cells. Thus, EBF1 and Pax5 may safeguard early stage B cells from transformation to B-ALL by limiting IL-7 signaling, folate metabolism and Myc expression.


Assuntos
Ácido Fólico/metabolismo , Interleucina-7/fisiologia , Fator de Transcrição PAX5/metabolismo , Leucemia-Linfoma Linfoblástico de Células Precursoras/fisiopatologia , Proteínas Proto-Oncogênicas c-myc/genética , Transdução de Sinais/genética , Transativadores/metabolismo , Animais , Carbono/metabolismo , Sobrevivência Celular/genética , Transformação Celular Neoplásica/genética , Modelos Animais de Doenças , Regulação Neoplásica da Expressão Gênica/genética , Redes Reguladoras de Genes/genética , Camundongos , Fator de Transcrição PAX5/genética , Leucemia-Linfoma Linfoblástico de Células Precursoras/genética , Células Precursoras de Linfócitos B/patologia , Ligação Proteica , Análise de Célula Única , Transativadores/genética
3.
Nature ; 572(7768): 199-204, 2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-31292543

RESUMO

The human liver is an essential multifunctional organ. The incidence of liver diseases is rising and there are limited treatment options. However, the cellular composition of the liver remains poorly understood. Here we performed single-cell RNA sequencing of about 10,000 cells from normal liver tissue from nine human donors to construct a human liver cell atlas. Our analysis identified previously unknown subtypes of endothelial cells, Kupffer cells, and hepatocytes, with transcriptome-wide zonation of some of these populations. We show that the EPCAM+ population is heterogeneous, comprising hepatocyte-biased and cholangiocyte populations as well as a TROP2int progenitor population with strong potential to form bipotent liver organoids. As a proof-of-principle, we used our atlas to unravel the phenotypic changes that occur in hepatocellular carcinoma cells and in human hepatocytes and liver endothelial cells engrafted into a mouse liver. Our human liver cell atlas provides a powerful resource to enable the discovery of previously unknown cell types in normal and diseased livers.


Assuntos
Células Epiteliais/citologia , Hepatócitos/citologia , Fígado/citologia , Células-Tronco/citologia , Adulto , Animais , Antígenos de Neoplasias/metabolismo , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/imunologia , Carcinoma Hepatocelular/patologia , Moléculas de Adesão Celular/metabolismo , Quimera/imunologia , Quimera/metabolismo , Células Endoteliais/citologia , Células Endoteliais/imunologia , Células Epiteliais/imunologia , Feminino , Regulação da Expressão Gênica , Hepatócitos/imunologia , Hepatócitos/metabolismo , Humanos , Fígado/imunologia , Masculino , Camundongos , Organoides/metabolismo , RNA Citoplasmático Pequeno/genética , RNA-Seq , Reprodutibilidade dos Testes , Células-Tronco/imunologia
4.
EMBO J ; 39(13): e104159, 2020 07 01.
Artigo em Inglês | MEDLINE | ID: mdl-32627520

RESUMO

γδ T cells with distinct properties develop in the embryonic and adult thymus and have been identified as critical players in a broad range of infections, antitumor surveillance, autoimmune diseases, and tissue homeostasis. Despite their potential value for immunotherapy, differentiation of γδ T cells in the thymus is incompletely understood. Here, we establish a high-resolution map of γδ T-cell differentiation from the fetal and adult thymus using single-cell RNA sequencing. We reveal novel sub-types of immature and mature γδ T cells and identify an unpolarized thymic population which is expanded in the blood and lymph nodes. Our detailed comparative analysis reveals remarkable similarities between the gene networks active during fetal and adult γδ T-cell differentiation. By performing a combined single-cell analysis of Sox13, Maf, and Rorc knockout mice, we demonstrate sequential activation of these factors during IL-17-producing γδ T-cell (γδT17) differentiation. These findings substantially expand our understanding of γδ T-cell ontogeny in fetal and adult life. Our experimental and computational strategy provides a blueprint for comparing immune cell differentiation across developmental stages.


Assuntos
Diferenciação Celular/imunologia , Feto/imunologia , Receptores de Antígenos de Linfócitos T gama-delta/imunologia , Linfócitos T/imunologia , Animais , Autoantígenos/genética , Autoantígenos/imunologia , Diferenciação Celular/genética , Camundongos , Camundongos Knockout , Membro 3 do Grupo F da Subfamília 1 de Receptores Nucleares/genética , Membro 3 do Grupo F da Subfamília 1 de Receptores Nucleares/imunologia , Proteínas Proto-Oncogênicas c-maf/genética , Proteínas Proto-Oncogênicas c-maf/imunologia , Receptores de Antígenos de Linfócitos T gama-delta/genética , Linfócitos T/citologia
5.
Nat Methods ; 15(5): 379-386, 2018 05.
Artigo em Inglês | MEDLINE | ID: mdl-29630061

RESUMO

To understand stem cell differentiation along multiple lineages, it is necessary to resolve heterogeneous cellular states and the ancestral relationships between them. We developed a robotic miniaturized CEL-Seq2 implementation to carry out deep single-cell RNA-seq of ∼2,000 mouse hematopoietic progenitors enriched for lymphoid lineages, and used an improved clustering algorithm, RaceID3, to identify cell types. To resolve subtle transcriptome differences indicative of lineage biases, we developed FateID, an iterative supervised learning algorithm for the probabilistic quantification of cell fate bias in progenitor populations. Here we used FateID to delineate domains of fate bias and enable the derivation of high-resolution differentiation trajectories, thereby revealing a common progenitor population of B cells and plasmacytoid dendritic cells, which we validated by in vitro differentiation assays. We expect that FateID will improve understanding of the process of cell fate choice in complex multi-lineage differentiation systems.


Assuntos
Diferenciação Celular/fisiologia , Células-Tronco Embrionárias/fisiologia , Software , Animais , Células da Medula Óssea , Técnicas de Cocultura , Biologia Computacional , Citometria de Fluxo , Regulação da Expressão Gênica , Masculino , Camundongos , Células-Tronco Multipotentes/fisiologia , Análise de Célula Única/métodos
6.
Cell Metab ; 34(4): 516-532.e11, 2022 04 05.
Artigo em Inglês | MEDLINE | ID: mdl-35316657

RESUMO

Metabolic reprogramming is a hallmark of activated T cells. The switch from oxidative phosphorylation to aerobic glycolysis provides energy and intermediary metabolites for the biosynthesis of macromolecules to support clonal expansion and effector function. Here, we show that glycolytic reprogramming additionally controls inflammatory gene expression via epigenetic remodeling. We found that the glucose transporter GLUT3 is essential for the effector functions of Th17 cells in models of autoimmune colitis and encephalomyelitis. At the molecular level, we show that GLUT3-dependent glucose uptake controls a metabolic-transcriptional circuit that regulates the pathogenicity of Th17 cells. Metabolomic, epigenetic, and transcriptomic analyses linked GLUT3 to mitochondrial glucose oxidation and ACLY-dependent acetyl-CoA generation as a rate-limiting step in the epigenetic regulation of inflammatory gene expression. Our findings are also important from a translational perspective because inhibiting GLUT3-dependent acetyl-CoA generation is a promising metabolic checkpoint to mitigate Th17-cell-mediated inflammatory diseases.


Assuntos
ATP Citrato (pro-S)-Liase , Transportador de Glucose Tipo 3 , Células Th17 , ATP Citrato (pro-S)-Liase/metabolismo , Acetilcoenzima A/metabolismo , Animais , Epigênese Genética , Glucose/metabolismo , Proteínas Facilitadoras de Transporte de Glucose/genética , Proteínas Facilitadoras de Transporte de Glucose/metabolismo , Transportador de Glucose Tipo 3/genética , Transportador de Glucose Tipo 3/metabolismo , Glicólise/genética , Humanos , Camundongos , Células Th17/metabolismo
7.
Nat Cell Biol ; 23(7): 704-717, 2021 07.
Artigo em Inglês | MEDLINE | ID: mdl-34253898

RESUMO

Haematopoietic stem cells (HSCs) are normally quiescent, but have evolved mechanisms to respond to stress. Here, we evaluate haematopoietic regeneration induced by chemotherapy. We detect robust chromatin reorganization followed by increased transcription of transposable elements (TEs) during early recovery. TE transcripts bind to and activate the innate immune receptor melanoma differentiation-associated protein 5 (MDA5) that generates an inflammatory response that is necessary for HSCs to exit quiescence. HSCs that lack MDA5 exhibit an impaired inflammatory response after chemotherapy and retain their quiescence, with consequent better long-term repopulation capacity. We show that the overexpression of ERV and LINE superfamily TE copies in wild-type HSCs, but not in Mda5-/- HSCs, results in their cycling. By contrast, after knockdown of LINE1 family copies, HSCs retain their quiescence. Our results show that TE transcripts act as ligands that activate MDA5 during haematopoietic regeneration, thereby enabling HSCs to mount an inflammatory response necessary for their exit from quiescence.


Assuntos
Proliferação de Células/efeitos dos fármacos , Senescência Celular/efeitos dos fármacos , Elementos de DNA Transponíveis , Hematopoese/efeitos dos fármacos , Células-Tronco Hematopoéticas/efeitos dos fármacos , Helicase IFIH1 Induzida por Interferon/metabolismo , Agonistas Mieloablativos/farmacologia , Animais , Montagem e Desmontagem da Cromatina/efeitos dos fármacos , Retrovirus Endógenos/genética , Ativação Enzimática , Células HEK293 , Células-Tronco Hematopoéticas/enzimologia , Humanos , Helicase IFIH1 Induzida por Interferon/genética , Ligantes , Elementos Nucleotídeos Longos e Dispersos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Transdução de Sinais
9.
Science ; 346(6213): 1109-13, 2014 Nov 28.
Artigo em Inglês | MEDLINE | ID: mdl-25378463

RESUMO

Mitochondria play central roles in cellular energy conversion, metabolism, and apoptosis. Mitochondria import more than 1000 different proteins from the cytosol. It is unknown if the mitochondrial protein import machinery is connected to the cell division cycle. We found that the cyclin-dependent kinase Cdk1 stimulated assembly of the main mitochondrial entry gate, the translocase of the outer membrane (TOM), in mitosis. The molecular mechanism involved phosphorylation of the cytosolic precursor of Tom6 by cyclin Clb3-activated Cdk1, leading to enhanced import of Tom6 into mitochondria. Tom6 phosphorylation promoted assembly of the protein import channel Tom40 and import of fusion proteins, thus stimulating the respiratory activity of mitochondria in mitosis. Tom6 phosphorylation provides a direct means for regulating mitochondrial biogenesis and activity in a cell cycle-specific manner.


Assuntos
Ciclo Celular , Mitocôndrias/metabolismo , Proteínas de Transporte da Membrana Mitocondrial/metabolismo , Precursores de Proteínas/metabolismo , Proteínas de Saccharomyces cerevisiae/metabolismo , Saccharomyces cerevisiae/citologia , Saccharomyces cerevisiae/metabolismo , Proteína Quinase CDC2/metabolismo , Ciclina B/metabolismo , Citosol/metabolismo , Proteínas do Complexo de Importação de Proteína Precursora Mitocondrial , Fosforilação , Transporte Proteico
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