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1.
Nat Med ; 27(1): 141-151, 2021 01.
Artigo em Inglês | MEDLINE | ID: mdl-33398161

RESUMO

Intratumoral heterogeneity (ITH) is a fundamental property of cancer; however, the origins of ITH remain poorly understood. We performed single-cell transcriptome profiling of peritoneal carcinomatosis (PC) from 15 patients with gastric adenocarcinoma (GAC), constructed a map of 45,048 PC cells, profiled the transcriptome states of tumor cell populations, incisively explored ITH of malignant PC cells and identified significant correlates with patient survival. The links between tumor cell lineage/state compositions and ITH were illustrated at transcriptomic, genotypic, molecular and phenotypic levels. We uncovered the diversity in tumor cell lineage/state compositions in PC specimens and defined it as a key contributor to ITH. Single-cell analysis of ITH classified PC specimens into two subtypes that were prognostically independent of clinical variables, and a 12-gene prognostic signature was derived and validated in multiple large-scale GAC cohorts. The prognostic signature appears fundamental to GAC carcinogenesis and progression and could be practical for patient stratification.


Assuntos
Adenocarcinoma/secundário , Neoplasias Gástricas/patologia , Adenocarcinoma/genética , Adenocarcinoma/patologia , Adulto , Idoso , Linhagem da Célula/genética , Cromossomos Humanos Par 17/genética , Estudos de Coortes , Variações do Número de Cópias de DNA , Feminino , Perfilação da Expressão Gênica , Variação Genética , Humanos , Masculino , Pessoa de Meia-Idade , Neoplasias Peritoneais/genética , Neoplasias Peritoneais/patologia , Neoplasias Peritoneais/secundário , Prognóstico , RNA-Seq , Análise de Célula Única , Neoplasias Gástricas/genética
2.
Nat Commun ; 12(1): 494, 2021 01 21.
Artigo em Inglês | MEDLINE | ID: mdl-33479210

RESUMO

Mast cells are critical effectors of allergic inflammation and protection against parasitic infections. We previously demonstrated that transcription factors GATA2 and MITF are the mast cell lineage-determining factors. However, it is unclear whether these lineage-determining factors regulate chromatin accessibility at mast cell enhancer regions. In this study, we demonstrate that GATA2 promotes chromatin accessibility at the super-enhancers of mast cell identity genes and primes both typical and super-enhancers at genes that respond to antigenic stimulation. We find that the number and densities of GATA2- but not MITF-bound sites at the super-enhancers are several folds higher than that at the typical enhancers. Our studies reveal that GATA2 promotes robust gene transcription to maintain mast cell identity and respond to antigenic stimulation by binding to super-enhancer regions with dense GATA2 binding sites available at key mast cell genes.


Assuntos
Antígenos/metabolismo , Montagem e Desmontagem da Cromatina/genética , Elementos Facilitadores Genéticos/genética , Fator de Transcrição GATA2/genética , Mastócitos/metabolismo , Animais , Antígenos/imunologia , Linhagem da Célula/genética , Células Cultivadas , Cromatina/genética , Cromatina/metabolismo , Feminino , Fator de Transcrição GATA2/metabolismo , Perfilação da Expressão Gênica/métodos , Masculino , Mastócitos/imunologia , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Transgênicos , Fator de Transcrição Associado à Microftalmia/genética , Fator de Transcrição Associado à Microftalmia/metabolismo
3.
Nat Commun ; 12(1): 652, 2021 01 28.
Artigo em Inglês | MEDLINE | ID: mdl-33510160

RESUMO

Injury and loss of oligodendrocytes can cause demyelinating diseases such as multiple sclerosis. To improve our understanding of human oligodendrocyte development, which could facilitate development of remyelination-based treatment strategies, here we describe time-course single-cell-transcriptomic analysis of developing human stem cell-derived oligodendrocyte-lineage-cells (hOLLCs). The study includes hOLLCs derived from both genome engineered embryonic stem cell (ESC) reporter cells containing an Identification-and-Purification tag driven by the endogenous PDGFRα promoter and from unmodified induced pluripotent (iPS) cells. Our analysis uncovers substantial transcriptional heterogeneity of PDGFRα-lineage hOLLCs. We discover sub-populations of human oligodendrocyte progenitor cells (hOPCs) including a potential cytokine-responsive hOPC subset, and identify candidate regulatory genes/networks that define the identity of these sub-populations. Pseudotime trajectory analysis defines developmental pathways of oligodendrocytes vs astrocytes from PDGFRα-expressing hOPCs and predicts differentially expressed genes between the two lineages. In addition, pathway enrichment analysis followed by pharmacological intervention of these pathways confirm that mTOR and cholesterol biosynthesis signaling pathways are involved in maturation of oligodendrocytes from hOPCs.


Assuntos
Heterogeneidade Genética , Variação Genética , Células-Tronco Pluripotentes Induzidas/metabolismo , Células Precursoras de Oligodendrócitos/metabolismo , Análise de Célula Única/métodos , Transcriptoma/genética , Astrócitos/citologia , Astrócitos/metabolismo , Diferenciação Celular/genética , Linhagem Celular , Linhagem da Célula/genética , Colesterol/biossíntese , Células-Tronco Embrionárias/citologia , Células-Tronco Embrionárias/metabolismo , Redes Reguladoras de Genes/genética , Humanos , Células-Tronco Pluripotentes Induzidas/citologia , Células Precursoras de Oligodendrócitos/citologia , Receptor alfa de Fator de Crescimento Derivado de Plaquetas/genética , Receptor alfa de Fator de Crescimento Derivado de Plaquetas/metabolismo , Transdução de Sinais/genética , Serina-Treonina Quinases TOR/genética , Serina-Treonina Quinases TOR/metabolismo
4.
Nat Commun ; 12(1): 420, 2021 01 18.
Artigo em Inglês | MEDLINE | ID: mdl-33462242

RESUMO

Adult stem cell identity, plasticity, and homeostasis are precisely orchestrated by lineage-restricted epigenetic and transcriptional regulatory networks. Here, by integrating super-enhancer and chromatin accessibility landscapes, we delineate core transcription regulatory circuitries (CRCs) of limbal stem/progenitor cells (LSCs) and find that RUNX1 and SMAD3 are required for maintenance of corneal epithelial identity and homeostasis. RUNX1 or SMAD3 depletion inhibits PAX6 and induces LSCs to differentiate into epidermal-like epithelial cells. RUNX1, PAX6, and SMAD3 (RPS) interact with each other and synergistically establish a CRC to govern the lineage-specific cis-regulatory atlas. Moreover, RUNX1 shapes LSC chromatin architecture via modulating H3K27ac deposition. Disturbance of RPS cooperation results in cell identity switching and dysfunction of the corneal epithelium, which is strongly linked to various human corneal diseases. Our work highlights CRC TF cooperativity for establishment of stem cell identity and lineage commitment, and provides comprehensive regulatory principles for human stratified epithelial homeostasis and pathogenesis.


Assuntos
Células-Tronco Adultas/metabolismo , Plasticidade Celular/genética , Doenças da Córnea/patologia , Epitélio Anterior/fisiologia , Redes Reguladoras de Genes/fisiologia , Adolescente , Adulto , Idoso , Linhagem da Célula/genética , Células Cultivadas , Criança , Cromatina/metabolismo , Sequenciamento de Cromatina por Imunoprecipitação , Subunidade alfa 2 de Fator de Ligação ao Core/genética , Subunidade alfa 2 de Fator de Ligação ao Core/metabolismo , Doenças da Córnea/genética , Epitélio Anterior/citologia , Feminino , Regulação da Expressão Gênica , Técnicas de Silenciamento de Genes , Humanos , Limbo da Córnea/citologia , Masculino , Pessoa de Meia-Idade , Fator de Transcrição PAX6/metabolismo , Cultura Primária de Células , RNA-Seq , Proteína Smad3/genética , Proteína Smad3/metabolismo
5.
Gene ; 764: 145101, 2021 Jan 05.
Artigo em Inglês | MEDLINE | ID: mdl-32877747

RESUMO

India is the world's largest milk producing country because of massive contribution made by cattle and buffaloes. In the present investigation, comprehensive comparative profiling of transcriptomic landscape of milk somatic cells of Sahiwal cattle and Murrah buffaloes was carried out. Genes with highest transcript abundance in both species were enriched for biological processes such as lactation, immune response, cellular oxidant detoxification and response to hormones. Analysis of differential expression identified 377 significantly up-regulated and 847 significantly down-regulated genes with fold change >1.5 in Murrah buffaloes as compared to Sahiwal cattle (padj <0.05). Marked enrichment of innate and adaptive immune response related GO terms and higher expression of genes for various host defense peptides such as lysozyme, defensin ß and granzymes were evident in buffaloes. Genes related to ECM-receptor interaction, complement and coagulation cascades, cytokine-cytokine receptor interaction and keratinization pathway showed more abundant expression in cattle. Network analysis of the up-regulated genes delineated highly connected genes representing immunity and haematopoietic cell lineage (CBL, CD28, CD247, PECAM1 and ITGA4). For the down-regulated dataset, genes with highest interactions were KRT18, FGFR1, GPR183, ITGB3 and DKK3. Our results lend support to more robust immune mechanisms in buffaloes, possibly explaining lower susceptibility to mammary infections as compared to cattle.


Assuntos
Búfalos/imunologia , Bovinos/imunologia , Imunidade/genética , Transcriptoma/imunologia , Animais , Peptídeos Catiônicos Antimicrobianos/genética , Búfalos/genética , Bovinos/genética , Linhagem da Célula/genética , Linhagem da Célula/imunologia , Regulação para Baixo/imunologia , Feminino , Hematopoese/genética , Hematopoese/imunologia , Índia , Lactação/genética , Lactação/imunologia , Leite/citologia , Leite/imunologia , RNA-Seq , Transcriptoma/genética , Regulação para Cima/imunologia
6.
Nat Metab ; 2(12): 1482-1497, 2020 12.
Artigo em Inglês | MEDLINE | ID: mdl-33324010

RESUMO

White and beige adipocytes in subcutaneous adipose tissue (SAT) and visceral adipose tissue (VAT) are maintained by proliferation and differentiation of adipose progenitor cells (APCs). Here we use mice with tissue-specific telomerase reverse transcriptase (TERT) gene knockout (KO), which undergo premature telomere shortening and proliferative senescence in APCs, to investigate the effect of over-nutrition on APC exhaustion and metabolic dysfunction. We find that TERT KO in the Pdgfra+ cell lineage results in adipocyte hypertrophy, inflammation and fibrosis in SAT, while TERT KO in the Pdgfrb+ lineage leads to adipocyte hypertrophy in both SAT and VAT. Systemic insulin resistance is observed in both KO models and is aggravated by a high-fat diet. Analysis of human biopsies demonstrates that telomere shortening in SAT is associated with metabolic disease progression after bariatric surgery. Our data indicate that over-nutrition can promote APC senescence and provide a mechanistic link between ageing, obesity and diabetes.


Assuntos
Adipócitos/patologia , Envelhecimento/patologia , Doenças Metabólicas/patologia , Células-Tronco/patologia , Homeostase do Telômero , Adipócitos Bege/metabolismo , Adipócitos Brancos/metabolismo , Animais , Diferenciação Celular , Linhagem da Célula/genética , Proliferação de Células , Dieta Hiperlipídica , Feminino , Humanos , Resistência à Insulina/genética , Gordura Intra-Abdominal , Masculino , Doenças Metabólicas/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Receptor alfa de Fator de Crescimento Derivado de Plaquetas/genética , Receptor alfa de Fator de Crescimento Derivado de Plaquetas/metabolismo , Gordura Subcutânea/metabolismo , Gordura Subcutânea/patologia , Telomerase/genética , Telomerase/metabolismo
7.
Nat Commun ; 11(1): 4483, 2020 09 08.
Artigo em Inglês | MEDLINE | ID: mdl-32900993

RESUMO

The Drosophila lymph gland, the larval hematopoietic organ comprised of prohemocytes and mature hemocytes, has been a valuable model for understanding mechanisms underlying hematopoiesis and immunity. Three types of mature hemocytes have been characterized in the lymph gland: plasmatocytes, lamellocytes, and crystal cells, which are analogous to vertebrate myeloid cells, yet molecular underpinnings of the lymph gland hemocytes have been less investigated. Here, we use single-cell RNA sequencing to comprehensively analyze heterogeneity of developing hemocytes in the lymph gland, and discover previously undescribed hemocyte types including adipohemocytes, stem-like prohemocytes, and intermediate prohemocytes. Additionally, we identify the developmental trajectory of hemocytes during normal development as well as the emergence of the lamellocyte lineage following active cellular immunity caused by wasp infestation. Finally, we establish similarities and differences between embryonically derived- and larval lymph gland hemocytes. Altogether, our study provides detailed insights into the hemocyte development and cellular immune responses at single-cell resolution.


Assuntos
Drosophila melanogaster/citologia , Drosophila melanogaster/genética , Hemócitos/citologia , Hemócitos/metabolismo , Transcriptoma , Animais , Animais Geneticamente Modificados , Diferenciação Celular/genética , Linhagem da Célula/genética , Drosophila melanogaster/metabolismo , Ectoparasitoses/genética , Ectoparasitoses/metabolismo , Ectoparasitoses/patologia , Perfilação da Expressão Gênica , Hematopoese/genética , Interações Hospedeiro-Parasita/genética , Interações Hospedeiro-Parasita/fisiologia , Tecido Linfoide/citologia , Tecido Linfoide/metabolismo , Tecido Linfoide/parasitologia , RNA-Seq , Análise de Célula Única , Vespas/patogenicidade
8.
Proc Natl Acad Sci U S A ; 117(40): 25043-25054, 2020 10 06.
Artigo em Inglês | MEDLINE | ID: mdl-32968015

RESUMO

Molecular and genomic surveillance systems for bacterial pathogens currently rely on tracking clonally evolving lineages. By contrast, plasmids are usually excluded or analyzed with low-resolution techniques, despite being the primary vectors of antibiotic resistance genes across many key pathogens. Here, we used a combination of long- and short-read sequence data of Klebsiella pneumoniae isolates (n = 1,717) from a European survey to perform an integrated, continent-wide study of chromosomal and plasmid diversity. This revealed three contrasting modes of dissemination used by carbapenemase genes, which confer resistance to last-line carbapenems. First, bla OXA-48-like genes have spread primarily via the single epidemic pOXA-48-like plasmid, which emerged recently in clinical settings and spread rapidly to numerous lineages. Second, bla VIM and bla NDM genes have spread via transient associations of many diverse plasmids with numerous lineages. Third, bla KPC genes have transmitted predominantly by stable association with one successful clonal lineage (ST258/512) yet have been mobilized among diverse plasmids within this lineage. We show that these plasmids, which include pKpQIL-like and IncX3 plasmids, have a long association (and are coevolving) with the lineage, although frequent recombination and rearrangement events between them have led to a complex array of mosaic plasmids carrying bla KPC Taken altogether, these results reveal the diverse trajectories of antibiotic resistance genes in clinical settings, summarized as using one plasmid/multiple lineages, multiple plasmids/multiple lineages, and multiple plasmids/one lineage. Our study provides a framework for the much needed incorporation of plasmid data into genomic surveillance systems, an essential step toward a more comprehensive understanding of resistance spread.


Assuntos
Proteínas de Bactérias/genética , Farmacorresistência Bacteriana Múltipla/genética , Infecções por Klebsiella/genética , Klebsiella pneumoniae/genética , beta-Lactamases/genética , Antibacterianos/uso terapêutico , Carbapenêmicos/uso terapêutico , Linhagem da Célula/genética , Cromossomos Bacterianos/genética , Farmacorresistência Bacteriana Múltipla/efeitos dos fármacos , Genoma Bacteriano/genética , Humanos , Infecções por Klebsiella/tratamento farmacológico , Infecções por Klebsiella/microbiologia , Klebsiella pneumoniae/patogenicidade , Plasmídeos/genética , Análise de Sequência de DNA/métodos
9.
Nat Commun ; 11(1): 4239, 2020 08 25.
Artigo em Inglês | MEDLINE | ID: mdl-32843640

RESUMO

How stem cells give rise to epidermis is unclear despite the crucial role the epidermis plays in barrier and appendage formation. Here we use single cell-RNA sequencing to interrogate basal stem cell heterogeneity of human interfollicular epidermis and find four spatially distinct stem cell populations at the top and bottom of rete ridges and transitional positions between the basal and suprabasal epidermal layers. Cell-cell communication modeling suggests that basal cell populations serve as crucial signaling hubs to maintain epidermal communication. Combining pseudotime, RNA velocity, and cellular entropy analyses point to a hierarchical differentiation lineage supporting multi-stem cell interfollicular epidermal homeostasis models and suggest that transitional basal stem cells are stable states essential for proper stratification. Finally, alterations in differentially expressed transitional basal stem cell genes result in severe thinning of human skin equivalents, validating their essential role in epidermal homeostasis and reinforcing the critical nature of basal stem cell heterogeneity.


Assuntos
Diferenciação Celular , Células Epidérmicas/citologia , Homeostase , Células-Tronco/citologia , Comunicação Celular/genética , Diferenciação Celular/genética , Linhagem da Célula/genética , Células Epidérmicas/metabolismo , Epiderme/metabolismo , Prepúcio do Pênis/citologia , Prepúcio do Pênis/metabolismo , Perfilação da Expressão Gênica , Regulação da Expressão Gênica , Humanos , Recém-Nascido , Queratinócitos/citologia , Queratinócitos/metabolismo , Masculino , Modelos Biológicos , Transdução de Sinais , Células-Tronco/metabolismo
10.
Nat Commun ; 11(1): 4158, 2020 08 27.
Artigo em Inglês | MEDLINE | ID: mdl-32855417

RESUMO

Visceral organs, such as the lungs, stomach and liver, are derived from the fetal foregut through a series of inductive interactions between the definitive endoderm (DE) and the surrounding splanchnic mesoderm (SM). While DE patterning is fairly well studied, the paracrine signaling controlling SM regionalization and how this is coordinated with epithelial identity is obscure. Here, we use single cell transcriptomics to generate a high-resolution cell state map of the embryonic mouse foregut. This identifies a diversity of SM cell types that develop in close register with the organ-specific epithelium. We infer a spatiotemporal signaling network of endoderm-mesoderm interactions that orchestrate foregut organogenesis. We validate key predictions with mouse genetics, showing the importance of endoderm-derived signals in mesoderm patterning. Finally, leveraging these signaling interactions, we generate different SM subtypes from human pluripotent stem cells (hPSCs), which previously have been elusive. The single cell data can be explored at: https://research.cchmc.org/ZornLab-singlecell .


Assuntos
Sistema Digestório/metabolismo , Endoderma/metabolismo , Redes Reguladoras de Genes , Mesoderma/metabolismo , Organogênese/genética , Transdução de Sinais/genética , Animais , Linhagem da Célula/genética , Sistema Digestório/citologia , Sistema Digestório/embriologia , Endoderma/citologia , Endoderma/embriologia , Perfilação da Expressão Gênica/métodos , Regulação da Expressão Gênica no Desenvolvimento , Humanos , Internet , Mesoderma/citologia , Mesoderma/embriologia , Camundongos Endogâmicos C57BL , Análise de Célula Única/métodos , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo
11.
Proc Natl Acad Sci U S A ; 117(34): 20729-20740, 2020 08 25.
Artigo em Inglês | MEDLINE | ID: mdl-32796104

RESUMO

Tissue-resident macrophages can originate from embryonic or adult hematopoiesis. They play important roles in a wide range of biological processes including tissue remodeling during organogenesis, organ homeostasis, repair following injury, and immune response to pathogens. Although the origins and tissue-specific functions of resident macrophages have been extensively studied in many other tissues, they are not well characterized in skeletal muscle. In the present study, we have characterized the ontogeny of skeletal muscle-resident macrophages by lineage tracing and bone marrow transplant experiments. We demonstrate that skeletal muscle-resident macrophages originate from both embryonic hematopoietic progenitors located within the yolk sac and fetal liver as well as definitive hematopoietic stem cells located within the bone marrow of adult mice. Single-cell-based transcriptome analyses revealed that skeletal muscle-resident macrophages are distinctive from resident macrophages in other tissues as they express a distinct complement of transcription factors and are composed of functionally diverse subsets correlating to their origins. Functionally, skeletal muscle-resident macrophages appear to maintain tissue homeostasis and promote muscle growth and regeneration.


Assuntos
Macrófagos/imunologia , Músculo Esquelético/imunologia , Animais , Medula Óssea/metabolismo , Transplante de Medula Óssea/métodos , Diferenciação Celular/genética , Linhagem da Célula/genética , Desenvolvimento Embrionário , Feminino , Heterogeneidade Genética , Hematopoese/genética , Células-Tronco Hematopoéticas/metabolismo , Homeostase , Macrófagos/metabolismo , Macrófagos/fisiologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Monócitos/imunologia , Monócitos/metabolismo , Organogênese/genética
12.
Nature ; 583(7818): 760-767, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32728245

RESUMO

During mammalian embryogenesis, differential gene expression gradually builds the identity and complexity of each tissue and organ system1. Here we systematically quantified mouse polyA-RNA from day 10.5 of embryonic development to birth, sampling 17 tissues and organs. The resulting developmental transcriptome is globally structured by dynamic cytodifferentiation, body-axis and cell-proliferation gene sets that were further characterized by the transcription factor motif codes of their promoters. We decomposed the tissue-level transcriptome using single-cell RNA-seq (sequencing of RNA reverse transcribed into cDNA) and found that neurogenesis and haematopoiesis dominate at both the gene and cellular levels, jointly accounting for one-third of differential gene expression and more than 40% of identified cell types. By integrating promoter sequence motifs with companion ENCODE epigenomic profiles, we identified a prominent promoter de-repression mechanism in neuronal expression clusters that was attributable to known and novel repressors. Focusing on the developing limb, single-cell RNA data identified 25 candidate cell types that included progenitor and differentiating states with computationally inferred lineage relationships. We extracted cell-type transcription factor networks and complementary sets of candidate enhancer elements by using single-cell RNA-seq to decompose integrative cis-element (IDEAS) models that were derived from whole-tissue epigenome chromatin data. These ENCODE reference data, computed network components and IDEAS chromatin segmentations are companion resources to the matching epigenomic developmental matrix, and are available for researchers to further mine and integrate.


Assuntos
Embrião de Mamíferos/citologia , Embrião de Mamíferos/embriologia , Desenvolvimento Embrionário/genética , Regulação da Expressão Gênica no Desenvolvimento , Análise de Célula Única , Transcriptoma , Animais , Diferenciação Celular/genética , Linhagem da Célula/genética , Cromatina/genética , Embrião de Mamíferos/metabolismo , Elementos Facilitadores Genéticos , Epigenômica , Extremidades/embriologia , Feminino , Masculino , Camundongos , Poli A/genética , Poli A/metabolismo , Regiões Promotoras Genéticas , RNA-Seq , Fatores de Transcrição/metabolismo
13.
Proc Natl Acad Sci U S A ; 117(29): 17041-17048, 2020 07 21.
Artigo em Inglês | MEDLINE | ID: mdl-32632001

RESUMO

A central task in developmental biology is to learn the sequence of fate decisions that leads to each mature cell type in a tissue or organism. Recently, clonal labeling of cells using DNA barcodes has emerged as a powerful approach for identifying cells that share a common ancestry of fate decisions. Here we explore the idea that stochasticity of cell fate choice during tissue development could be harnessed to read out lineage relationships after a single step of clonal barcoding. By considering a generalized multitype branching process, we determine the conditions under which the final distribution of barcodes over observed cell types encodes their bona fide lineage relationships. We then propose a method for inferring the order of fate decisions. Our theory predicts a set of symmetries of barcode covariance that serves as a consistency check for the validity of the method. We show that broken symmetries may be used to detect multiple paths of differentiation to the same cell types. We provide computational tools for general use. When applied to barcoding data in hematopoiesis, these tools reconstruct the classical hematopoietic hierarchy and detect couplings between monocytes and dendritic cells and between erythrocytes and basophils that suggest multiple pathways of differentiation for these lineages.


Assuntos
Linhagem da Célula , Código de Barras de DNA Taxonômico/métodos , Animais , Linhagem da Célula/genética , Linhagem da Célula/fisiologia , Árvores de Decisões , Células Dendríticas/citologia , Eritrócitos/citologia , Hematopoese/genética , Hematopoese/fisiologia , Leucócitos/citologia , Modelos Biológicos , Biologia de Sistemas
15.
Nat Med ; 26(8): 1295-1306, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32601335

RESUMO

Immune-regulatory mechanisms of drug-free remission in rheumatoid arthritis (RA) are unknown. We hypothesized that synovial tissue macrophages (STM), which persist in remission, contribute to joint homeostasis. We used single-cell transcriptomics to profile 32,000 STMs and identified phenotypic changes in patients with early/active RA, treatment-refractory/active RA and RA in sustained remission. Each clinical state was characterized by different frequencies of nine discrete phenotypic clusters within four distinct STM subpopulations with diverse homeostatic, regulatory and inflammatory functions. This cellular atlas, combined with deep-phenotypic, spatial and functional analyses of synovial biopsy fluorescent activated cell sorted STMs, revealed two STM subpopulations (MerTKposTREM2high and MerTKposLYVE1pos) with unique remission transcriptomic signatures enriched in negative regulators of inflammation. These STMs were potent producers of inflammation-resolving lipid mediators and induced the repair response of synovial fibroblasts in vitro. A low proportion of MerTKpos STMs in remission was associated with increased risk of disease flare after treatment cessation. Therapeutic modulation of MerTKpos STM subpopulations could therefore be a potential treatment strategy for RA.


Assuntos
Artrite Reumatoide/metabolismo , Inflamação/metabolismo , Macrófagos/imunologia , Líquido Sinovial/metabolismo , Artrite Reumatoide/genética , Artrite Reumatoide/imunologia , Artrite Reumatoide/patologia , Biópsia , Linhagem da Célula/genética , Humanos , Inflamação/genética , Inflamação/imunologia , Inflamação/patologia , Articulações/imunologia , Articulações/metabolismo , Articulações/patologia , Lectinas Tipo C/genética , Lectinas Tipo C/imunologia , Macrófagos/metabolismo , Lectinas de Ligação a Manose/genética , Lectinas de Ligação a Manose/imunologia , Receptores de Superfície Celular/genética , Receptores de Superfície Celular/imunologia , Receptores Imunológicos/genética , Receptores Imunológicos/imunologia , Líquido Sinovial/imunologia , Membrana Sinovial
16.
Nat Cell Biol ; 22(8): 919-926, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32690888

RESUMO

Intestinal stem cells (ISCs) are located at the crypt base and fine-tune the balance of their self-renewal and differentiation1,2, but the physiological mechanism involved in regulating that balance remains unknown. Here we describe a transcriptional regulator that preserves the stemness of ISCs by restricting their differentiation into secretory-cell lineages. Interferon regulatory factor 2 (IRF2) negatively regulates interferon signalling3, and mice completely lacking Irf24 or with a selective Irf2 deletion in their intestinal epithelial cells have significantly fewer crypt Lgr5hi ISCs than control mice. Although the integrity of intestinal epithelial cells was unimpaired at steady state in Irf2-deficient mice, regeneration of their intestinal epithelia after 5-fluorouracil-induced damage was severely impaired. Similarly, extended treatment with low-dose poly(I:C) or chronic infection of lymphocytic choriomeningitis virus clone 13 (LCMV C13)5 caused a functional decline of ISCs in wild-type mice. In contrast, massive accumulations of immature Paneth cells were found at the crypt base of Irf2-/- as well as LCMV C13-infected wild-type mice, indicating that excess interferon signalling directs ISCs towards a secretory-cell fate. Collectively, our findings indicate that regulated interferon signalling preserves ISC stemness by restricting secretory-cell differentiation.


Assuntos
Linhagem da Célula , Fator Regulador 2 de Interferon/metabolismo , Mucosa Intestinal/citologia , Transdução de Sinais , Células-Tronco/metabolismo , Idoso , Animais , Diferenciação Celular/genética , Linhagem da Célula/genética , Feminino , Regulação da Expressão Gênica , Humanos , Interferons/metabolismo , Mucosa Intestinal/metabolismo , Secreções Intestinais , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Pessoa de Meia-Idade , Células-Tronco/citologia
17.
PLoS Comput Biol ; 16(7): e1007731, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32649725

RESUMO

High-throughput sequencing of human immunoglobulin genes allows analysis of antibody repertoires and the reconstruction of clonal lineage evolution. The study of antibodies (Abs) affinity maturation is of specific interest to understand the generation of Abs with high affinity or broadly neutralizing activities. Moreover, phylogenic analysis enables the identification of the key somatic mutations required to achieve optimal antigen binding. The Immcantation framework provides a start-to-finish set of analytical methods for high-throughput adaptive immune receptor repertoire sequencing (AIRR-Seq; Rep-Seq) data. Furthermore, Immcantation's Change-O package has developed IgPhyML, an algorithm designed to build specifically immunoglobulin (Ig) phylogenic trees. Meanwhile Phylip, an algorithm that has been originally developed for applications in ecology and macroevolution, can also be used for the phylogenic reconstruction of antibodies maturation pathway. To complement Ig lineages made by IgPhyML or Dnaml (Phylip), we developed AncesTree, a graphic user interface (GUI) that aims to give researchers the opportunity to interactively explore antibodies clonal evolution. AncesTree displays interactive immunoglobulins phylogenic tree, Ig related mutations and sequence alignments using additional information coming from specialized antibody tools. The GUI is a Java standalone application allowing interaction with Ig tree that can run under Windows, Linux and Mac OS.


Assuntos
Genes de Imunoglobulinas/genética , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Imunoglobulinas , Alinhamento de Sequência/métodos , Software , Algoritmos , Linhagem da Célula/genética , Humanos , Imunoglobulinas/química , Imunoglobulinas/classificação , Imunoglobulinas/genética , Filogenia , Análise de Sequência de DNA/métodos
18.
Nat Commun ; 11(1): 3055, 2020 06 16.
Artigo em Inglês | MEDLINE | ID: mdl-32546686

RESUMO

Recent studies combine two novel technologies, single-cell RNA-sequencing and CRISPR-Cas9 barcode editing for elucidating developmental lineages at the whole organism level. While these studies provided several insights, they face several computational challenges. First, lineages are reconstructed based on noisy and often saturated random mutation data. Additionally, due to the randomness of the mutations, lineages from multiple experiments cannot be combined to reconstruct a species-invariant lineage tree. To address these issues we developed a statistical method, LinTIMaT, which reconstructs cell lineages using a maximum-likelihood framework by integrating mutation and expression data. Our analysis shows that expression data helps resolve the ambiguities arising in when lineages are inferred based on mutations alone, while also enabling the integration of different individual lineages for the reconstruction of an invariant lineage tree. LinTIMaT lineages have better cell type coherence, improve the functional significance of gene sets and provide new insights on progenitors and differentiation pathways.


Assuntos
Algoritmos , Sistemas CRISPR-Cas , Linhagem da Célula/genética , Perfilação da Expressão Gênica/estatística & dados numéricos , Análise de Célula Única/métodos , Animais , Encéfalo/citologia , Caenorhabditis elegans/embriologia , Caenorhabditis elegans/genética , Diferenciação Celular/genética , Interpretação Estatística de Dados , Embrião não Mamífero/citologia , Perfilação da Expressão Gênica/métodos , Funções Verossimilhança , Mutação , Análise de Célula Única/estatística & dados numéricos , Peixe-Zebra/genética
19.
Nat Commun ; 11(1): 3013, 2020 06 15.
Artigo em Inglês | MEDLINE | ID: mdl-32541654

RESUMO

B lymphoid development is initiated by the differentiation of hematopoietic stem cells into lineage committed progenitors, ultimately generating mature B cells. This highly regulated process generates clonal immunological diversity via recombination of immunoglobulin V, D and J gene segments. While several transcription factors that control B cell development and V(D)J recombination have been defined, how these processes are initiated and coordinated into a precise regulatory network remains poorly understood. Here, we show that the transcription factor ETS Related Gene (Erg) is essential for early B lymphoid differentiation. Erg initiates a transcriptional network involving the B cell lineage defining genes, Ebf1 and Pax5, which directly promotes expression of key genes involved in V(D)J recombination and formation of the B cell receptor. Complementation of Erg deficiency with a productively rearranged immunoglobulin gene rescued B lineage development, demonstrating that Erg is an essential and stage-specific regulator of the gene regulatory network controlling B lymphopoiesis.


Assuntos
Linfócitos B/metabolismo , Diferenciação Celular/genética , Células-Tronco Hematopoéticas/metabolismo , Linfopoese/genética , Proteínas Oncogênicas/genética , Transcrição Genética , Regulador Transcricional ERG/genética , Animais , Linfócitos B/citologia , Linhagem da Célula/genética , Células Cultivadas , Redes Reguladoras de Genes/genética , Células-Tronco Hematopoéticas/citologia , Camundongos Endogâmicos C57BL , Camundongos Knockout , Proteínas Oncogênicas/metabolismo , Fator de Transcrição PAX5/genética , Fator de Transcrição PAX5/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Regulador Transcricional ERG/metabolismo , Recombinação V(D)J/genética
20.
Nature ; 582(7813): 534-538, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32555454

RESUMO

Many corals harbour symbiotic dinoflagellate algae. The algae live inside coral cells in a specialized membrane compartment known as the symbiosome, which shares the photosynthetically fixed carbon with coral host cells while host cells provide inorganic carbon to the algae for photosynthesis1. This endosymbiosis-which is critical for the maintenance of coral reef ecosystems-is increasingly threatened by environmental stressors that lead to coral bleaching (that is, the disruption of endosymbiosis), which in turn leads to coral death and the degradation of marine ecosystems2. The molecular pathways that orchestrate the recognition, uptake and maintenance of algae in coral cells remain poorly understood. Here we report the chromosome-level genome assembly of a Xenia species of fast-growing soft coral3, and use this species as a model to investigate coral-alga endosymbiosis. Single-cell RNA sequencing identified 16 cell clusters, including gastrodermal cells and cnidocytes, in Xenia sp. We identified the endosymbiotic cell type, which expresses a distinct set of genes that are implicated in the recognition, phagocytosis and/or endocytosis, and maintenance of algae, as well as in the immune modulation of host coral cells. By coupling Xenia sp. regeneration and single-cell RNA sequencing, we observed a dynamic lineage progression of the endosymbiotic cells. The conserved genes associated with endosymbiosis that are reported here may help to reveal common principles by which different corals take up or lose their endosymbionts.


Assuntos
Antozoários/citologia , Antozoários/genética , Linhagem da Célula/genética , Dinoflagelados/metabolismo , Simbiose/genética , Animais , Antozoários/imunologia , Antozoários/metabolismo , Carbono/metabolismo , Diferenciação Celular/genética , Recifes de Corais , Dinoflagelados/imunologia , Dinoflagelados/fisiologia , Ecossistema , Endocitose , Genoma/genética , Fagocitose , Fotossíntese , RNA-Seq , Análise de Célula Única , Simbiose/imunologia , Transcriptoma
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