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1.
Nat Commun ; 12(1): 4693, 2021 08 03.
Artigo em Inglês | MEDLINE | ID: mdl-34344862

RESUMO

Many cellular processes, including cell division, development, and cell migration require spatially and temporally coordinated forces transduced by cell-surface receptors. Nucleic acid-based molecular tension probes allow one to visualize the piconewton (pN) forces applied by these receptors. Building on this technology, we recently developed molecular force microscopy (MFM) which uses fluorescence polarization to map receptor force orientation with diffraction-limited resolution (~250 nm). Here, we show that structured illumination microscopy (SIM), a super-resolution technique, can be used to perform super-resolution MFM. Using SIM-MFM, we generate the highest resolution maps of both the magnitude and orientation of the pN traction forces applied by cells. We apply SIM-MFM to map platelet and fibroblast integrin forces, as well as T cell receptor forces. Using SIM-MFM, we show that platelet traction force alignment occurs on a longer timescale than adhesion. Importantly, SIM-MFM can be implemented on any standard SIM microscope without hardware modifications.


Assuntos
Microscopia de Fluorescência/métodos , Receptores de Superfície Celular/metabolismo , Animais , Fenômenos Biomecânicos , Plaquetas/metabolismo , Linfócitos T CD8-Positivos , Corantes Fluorescentes/metabolismo , Humanos , Integrinas/metabolismo , Camundongos , Sondas Moleculares/metabolismo , Células NIH 3T3 , Paxilina/metabolismo , Receptores de Antígenos de Linfócitos T/metabolismo , Imagem com Lapso de Tempo
2.
Nat Commun ; 12(1): 4515, 2021 07 26.
Artigo em Inglês | MEDLINE | ID: mdl-34312385

RESUMO

The in vivo phenotypic profile of T cells reactive to severe acute respiratory syndrome (SARS)-CoV-2 antigens remains poorly understood. Conventional methods to detect antigen-reactive T cells require in vitro antigenic re-stimulation or highly individualized peptide-human leukocyte antigen (pHLA) multimers. Here, we use single-cell RNA sequencing to identify and profile SARS-CoV-2-reactive T cells from Coronavirus Disease 2019 (COVID-19) patients. To do so, we induce transcriptional shifts by antigenic stimulation in vitro and take advantage of natural T cell receptor (TCR) sequences of clonally expanded T cells as barcodes for 'reverse phenotyping'. This allows identification of SARS-CoV-2-reactive TCRs and reveals phenotypic effects introduced by antigen-specific stimulation. We characterize transcriptional signatures of currently and previously activated SARS-CoV-2-reactive T cells, and show correspondence with phenotypes of T cells from the respiratory tract of patients with severe disease in the presence or absence of virus in independent cohorts. Reverse phenotyping is a powerful tool to provide an integrated insight into cellular states of SARS-CoV-2-reactive T cells across tissues and activation states.


Assuntos
COVID-19/imunologia , Perfilação da Expressão Gênica/métodos , Análise de Sequência de RNA/métodos , Análise de Célula Única/métodos , Linfócitos T/metabolismo , Idoso , Idoso de 80 Anos ou mais , Linfócitos T CD4-Positivos/metabolismo , Linfócitos T CD4-Positivos/virologia , COVID-19/epidemiologia , COVID-19/virologia , Células Cultivadas , Estudos de Coortes , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Pandemias , Receptores de Antígenos de Linfócitos T/genética , Receptores de Antígenos de Linfócitos T/imunologia , Receptores de Antígenos de Linfócitos T/metabolismo , SARS-CoV-2/fisiologia , Linfócitos T/virologia
3.
Nat Immunol ; 22(7): 809-819, 2021 07.
Artigo em Inglês | MEDLINE | ID: mdl-34140679

RESUMO

CD8+ T cells are critical mediators of cytotoxic effector function in infection, cancer and autoimmunity. In cancer and chronic viral infection, CD8+ T cells undergo a progressive loss of cytokine production and cytotoxicity, a state termed T cell exhaustion. In autoimmunity, autoreactive CD8+ T cells retain the capacity to effectively mediate the destruction of host tissues. Although the clinical outcome differs in each context, CD8+ T cells are chronically exposed to antigen in all three. These chronically stimulated CD8+ T cells share some common phenotypic features, as well as transcriptional and epigenetic programming, across disease contexts. A better understanding of these CD8+ T cell states may reveal novel strategies to augment clearance of chronic viral infection and cancer and to mitigate self-reactivity leading to tissue damage in autoimmunity.


Assuntos
Doenças Autoimunes/imunologia , Autoimunidade , Linfócitos T CD8-Positivos/imunologia , Doenças Transmissíveis/imunologia , Linfócitos do Interstício Tumoral/imunologia , Neoplasias/imunologia , Animais , Doenças Autoimunes/genética , Doenças Autoimunes/metabolismo , Antígeno B7-H1/imunologia , Antígeno B7-H1/metabolismo , Linfócitos T CD8-Positivos/efeitos dos fármacos , Linfócitos T CD8-Positivos/metabolismo , Doença Crônica , Doenças Transmissíveis/genética , Doenças Transmissíveis/metabolismo , Citocinas/imunologia , Citocinas/metabolismo , Epigênese Genética , Humanos , Inibidores de Checkpoint Imunológico/uso terapêutico , Linfócitos do Interstício Tumoral/efeitos dos fármacos , Linfócitos do Interstício Tumoral/metabolismo , Neoplasias/tratamento farmacológico , Neoplasias/genética , Neoplasias/metabolismo , Fenótipo , Receptor de Morte Celular Programada 1/imunologia , Receptor de Morte Celular Programada 1/metabolismo , Receptores de Antígenos de Linfócitos T/imunologia , Receptores de Antígenos de Linfócitos T/metabolismo , Transdução de Sinais
4.
Nat Commun ; 12(1): 3933, 2021 06 24.
Artigo em Inglês | MEDLINE | ID: mdl-34168132

RESUMO

Thymic T cell development and T cell receptor repertoire selection are dependent on essential molecular cues provided by thymic epithelial cells (TEC). TEC development and function are regulated by their epigenetic landscape, in which the repressive H3K27me3 epigenetic marks are catalyzed by polycomb repressive complex 2 (PRC2). Here we show that a TEC-targeted deficiency of PRC2 function results in a hypoplastic thymus with reduced ability to express antigens and select a normal repertoire of T cells. The absence of PRC2 activity reveals a transcriptomically distinct medullary TEC lineage that incompletely off-sets the shortage of canonically-derived medullary TEC whereas cortical TEC numbers remain unchanged. This alternative TEC development is associated with the generation of reduced TCR diversity. Hence, normal PRC2 activity and placement of H3K27me3 marks are required for TEC lineage differentiation and function and, in their absence, the thymus is unable to compensate for the loss of a normal TEC scaffold.


Assuntos
Epigênese Genética , Células Epiteliais/citologia , Complexo Repressor Polycomb 2/genética , Timo/citologia , Animais , Linfócitos T CD4-Positivos/metabolismo , Linfócitos T CD8-Positivos/metabolismo , Diferenciação Celular , Linhagem da Célula , Células Epiteliais/fisiologia , Feminino , Masculino , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Complexo Repressor Polycomb 2/metabolismo , Receptores de Antígenos de Linfócitos T/metabolismo , Linfócitos T/citologia , Linfócitos T/fisiologia , Timócitos/citologia , Timócitos/fisiologia , Timo/fisiologia
5.
Eur J Immunol ; 51(8): 1992-2005, 2021 08.
Artigo em Inglês | MEDLINE | ID: mdl-34081326

RESUMO

The phenotype of infused cells is a major determinant of Adoptive T-cell therapy (ACT) efficacy. Yet, the difficulty in deciphering multiparametric cytometry data limited the fine characterization of cellular products. To allow the analysis of dynamic and complex flow cytometry samples, we developed cytoChain, a novel dataset mining tool and a new analytical workflow. CytoChain was challenged to compare state-of-the-art and innovative culture conditions to generate stem-like memory cells (TSCM ) suitable for ACT. Noticeably, the combination of IL-7/15 and superoxides scavenging sustained the emergence of a previously unidentified nonexhausted Fit-TSCM signature, overlooked by manual gating and endowed with superior expansion potential. CytoChain proficiently traced back this population in independent datasets, and in T-cell receptor engineered lymphocytes. CytoChain flexibility and function were then further validated on a published dataset from circulating T cells in COVID-19 patients. Collectively, our results support the use of cytoChain to identify novel, functionally critical immunophenotypes for ACT and patients immunomonitoring.


Assuntos
Mineração de Dados/métodos , Citometria de Fluxo/métodos , Receptores de Antígenos de Linfócitos T/imunologia , Receptores de Antígenos Quiméricos/imunologia , Linfócitos T/imunologia , Linfócitos T/metabolismo , COVID-19/sangue , COVID-19/imunologia , Citocinas/metabolismo , Engenharia Genética , Humanos , Memória Imunológica , Imunofenotipagem , Imunoterapia Adotiva , Receptores de Antígenos de Linfócitos T/genética , Receptores de Antígenos de Linfócitos T/metabolismo , Receptores de Antígenos Quiméricos/genética , SARS-CoV-2/imunologia
6.
Nat Commun ; 12(1): 3705, 2021 06 17.
Artigo em Inglês | MEDLINE | ID: mdl-34140493

RESUMO

Peripheral T-cell lymphoma (PTCL) is a heterogeneous group of non-Hodgkin lymphomas with poor prognosis. Up to 30% of PTCL lack distinctive features and are classified as PTCL, not otherwise specified (PTCL-NOS). To further improve our understanding of the genetic landscape and biology of PTCL-NOS, we perform RNA-sequencing of 18 cases and validate results in an independent cohort of 37 PTCL cases. We identify FYN-TRAF3IP2, KHDRBS1-LCK and SIN3A-FOXO1 as new in-frame fusion transcripts, with FYN-TRAF3IP2 as a recurrent fusion detected in 8 of 55 cases. Using ex vivo and in vivo experiments, we demonstrate that FYN-TRAF3IP2 and KHDRBS1-LCK activate signaling pathways downstream of the T cell receptor (TCR) complex and confer therapeutic vulnerability to clinically available drugs.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas de Ligação a DNA/genética , Proteína Tirosina Quinase p56(lck) Linfócito-Específica/genética , Linfoma de Células T Periférico/genética , Proteínas de Fusão Oncogênica/metabolismo , Proteínas Proto-Oncogênicas c-fyn/genética , Proteínas de Ligação a RNA/genética , Receptores de Antígenos de Linfócitos T/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Animais , Linhagem Celular Tumoral , Membrana Celular/metabolismo , Estudos de Coortes , Proteínas de Ligação a DNA/metabolismo , Proteína Forkhead Box O1/genética , Proteína Forkhead Box O1/metabolismo , Regulação Neoplásica da Expressão Gênica/genética , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Estimativa de Kaplan-Meier , Proteína Tirosina Quinase p56(lck) Linfócito-Específica/metabolismo , Linfoma de Células T Periférico/metabolismo , Linfoma de Células T Periférico/patologia , Camundongos , Camundongos Endogâmicos C57BL , NF-kappa B/metabolismo , Proteínas de Fusão Oncogênica/genética , Proteínas Proto-Oncogênicas c-fyn/metabolismo , Proteínas de Ligação a RNA/metabolismo , RNA-Seq , Transdução de Sinais/genética , Complexo Correpressor Histona Desacetilase e Sin3/genética , Complexo Correpressor Histona Desacetilase e Sin3/metabolismo , Proteína bcl-X/antagonistas & inibidores , Proteína bcl-X/metabolismo
7.
Nat Commun ; 12(1): 3615, 2021 06 14.
Artigo em Inglês | MEDLINE | ID: mdl-34127674

RESUMO

Glioblastoma is considered one of the most aggressive malignancies in adult and pediatric patients. Despite decades of research no curative treatment is available and it thus remains associated with a very dismal prognosis. Although recent pre-clinical and clinical studies have demonstrated the feasibility of chimeric antigen receptors (CAR) T cell immunotherapeutic approach in glioblastoma, tumor heterogeneity and antigen loss remain among one of the most important challenges to be addressed. In this study, we identify p32/gC1qR/HABP/C1qBP to be specifically expressed on the surface of glioma cells, making it a suitable tumor associated antigen for redirected CAR T cell therapy. We generate p32 CAR T cells and find them to recognize and specifically eliminate p32 expressing glioma cells and tumor derived endothelial cells in vitro and to control tumor growth in orthotopic syngeneic and xenograft mouse models. Thus, p32 CAR T cells may serve as a therapeutic option for glioblastoma patients.


Assuntos
Inibidores da Angiogênese/uso terapêutico , Antineoplásicos/farmacologia , Glioma/imunologia , Glioma/terapia , Linfócitos T/imunologia , Idoso , Animais , Antígenos de Neoplasias/imunologia , Neoplasias Encefálicas , Proteínas de Transporte/metabolismo , Linhagem Celular Tumoral , Feminino , Glioblastoma/genética , Glioblastoma/patologia , Glioma/genética , Glioma/metabolismo , Humanos , Imunoterapia Adotiva , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Nus , Proteínas Mitocondriais/metabolismo , Receptores de Antígenos de Linfócitos T/metabolismo , Receptores de Antígenos Quiméricos/imunologia , Serina Endopeptidases/metabolismo , Ensaios Antitumorais Modelo de Xenoenxerto
8.
Front Immunol ; 12: 616967, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34108957

RESUMO

The function of mucosal-associated invariant T (MAIT) cells highly depends on the mode of activation, either by recognition of bacterial metabolites via their T cell receptor (TCR) or in a TCR-independent manner via cytokines. The underlying molecular mechanisms are not entirely understood. To define the activation of MAIT cells on the molecular level, we applied a multi-omics approach with untargeted transcriptomics, proteomics and metabolomics. Transcriptomic analysis of E. coli- and TCR-activated MAIT cells showed a distinct transcriptional reprogramming, including altered pathways, transcription factors and effector molecules. We validated the consequences of this reprogramming on the phenotype by proteomics and metabolomics. Thus, and to distinguish between TCR-dependent and -independent activation, MAIT cells were stimulated with IL12/IL18, anti-CD3/CD28 or both. Only a combination of both led to full activation of MAIT cells, comparable to activation by E. coli. Using an integrated network-based approach, we identified key drivers of the distinct modes of activation, including cytokines and transcription factors, as well as negative feedback regulators like TWIST1 or LAG3. Taken together, we present novel insights into the biological function of MAIT cells, which may represent a basis for therapeutic approaches to target MAIT cells in pathological conditions.


Assuntos
Perfilação da Expressão Gênica , Ativação Linfocitária/genética , Ativação Linfocitária/imunologia , Metabolômica , Células T Invariantes Associadas à Mucosa/imunologia , Células T Invariantes Associadas à Mucosa/metabolismo , Proteômica , Biomarcadores , Células Cultivadas , Cromatografia Líquida , Biologia Computacional/métodos , Perfilação da Expressão Gênica/métodos , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Imunofenotipagem , Metabolômica/métodos , Proteômica/métodos , Receptores de Antígenos de Linfócitos T/metabolismo , Espectrometria de Massas em Tandem
9.
Int J Mol Sci ; 22(9)2021 May 06.
Artigo em Inglês | MEDLINE | ID: mdl-34066527

RESUMO

Activation of T cells by agonistic peptide-MHC can be inhibited by antagonistic ones. However, the exact mechanism remains elusive. We used Jurkat cells expressing two different TCRs and tested whether stimulation of the endogenous TCR by agonistic anti-Vß8 antibodies can be modulated by ligand-binding to the second, optogenetic TCR. The latter TCR uses phytochrome B tetramers (PhyBt) as ligand, the binding half-life of which can be altered by light. We show that this half-life determined whether the PhyBt acted as a second agonist (long half-life), an antagonist (short half-life) or did not have any influence (very short half-life) on calcium influx. A mathematical model of this cross-antagonism shows that a mechanism based on an inhibitory signal generated by early recruitment of a phosphatase and an activating signal by later recruitment of a kinase explains the data.


Assuntos
Optogenética , Receptores de Antígenos de Linfócitos T/antagonistas & inibidores , Anticorpos/metabolismo , Membrana Celular/metabolismo , Proteínas de Fluorescência Verde/metabolismo , Células HEK293 , Meia-Vida , Humanos , Células Jurkat , Ligantes , Modelos Biológicos , Receptores de Antígenos de Linfócitos T/metabolismo
10.
Nat Commun ; 12(1): 3872, 2021 06 23.
Artigo em Inglês | MEDLINE | ID: mdl-34162836

RESUMO

The tyrosine phosphatase CD45 is a major gatekeeper for restraining T cell activation. Its exclusion from the immunological synapse (IS) is crucial for T cell receptor (TCR) signal transduction. Here, we use expansion super-resolution microscopy to reveal that CD45 is mostly pre-excluded from the tips of microvilli (MV) on primary T cells prior to antigen encounter. This pre-exclusion is diminished by depleting cholesterol or by engineering the transmembrane domain of CD45 to increase its membrane integration length, but is independent of the CD45 extracellular domain. We further show that brief MV-mediated contacts can induce Ca2+ influx in mouse antigen-specific T cells engaged by antigen-pulsed antigen presenting cells (APC). We propose that the scarcity of CD45 phosphatase activity at the tips of MV enables or facilitates TCR triggering from brief T cell-APC contacts before formation of a stable IS, and that these MV-mediated contacts represent the earliest step in the initiation of a T cell adaptive immune response.


Assuntos
Células Apresentadoras de Antígenos/imunologia , Antígenos Comuns de Leucócito/imunologia , Microvilosidades/imunologia , Receptores de Antígenos de Linfócitos T/imunologia , Linfócitos T/imunologia , Animais , Células Apresentadoras de Antígenos/metabolismo , Células Cultivadas , Feminino , Células HEK293 , Humanos , Células Jurkat , Antígenos Comuns de Leucócito/genética , Antígenos Comuns de Leucócito/metabolismo , Ativação Linfocitária/imunologia , Masculino , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Transgênicos , Microvilosidades/metabolismo , Fosforilação/imunologia , Proteínas Tirosina Fosfatases/genética , Proteínas Tirosina Fosfatases/imunologia , Proteínas Tirosina Fosfatases/metabolismo , Receptores de Antígenos de Linfócitos T/metabolismo , Transdução de Sinais/imunologia , Linfócitos T/metabolismo
11.
J Virol ; 95(14): e0162820, 2021 06 24.
Artigo em Inglês | MEDLINE | ID: mdl-33952641

RESUMO

Like all herpesviruses, the roseoloviruses (HHV6A, -6B, and -7) establish lifelong infection within their host, requiring these viruses to evade host antiviral responses. One common host-evasion strategy is the downregulation of host-encoded, surface-expressed glycoproteins. Roseoloviruses have been shown to evade the host immune response by downregulating NK-activating ligands, class I MHC, and the TCR/CD3 complex. To more globally identify glycoproteins that are differentially expressed on the surface of HHV6A-infected cells, we performed cell surface capture of N-linked glycoproteins present on the surface of T cells infected with HHV6A, and compared these to proteins present on the surface of uninfected T cells. We found that the protein tyrosine phosphatase CD45 is downregulated in T cells infected with HHV6A. We also demonstrated that CD45 is similarly downregulated in cells infected with HHV7. CD45 is essential for signaling through the T cell receptor and, as such, is necessary for developing a fully functional immune response. Interestingly, the closely related betaherpesviruses human cytomegalovirus (HCMV) and murine cytomegalovirus (MCMV) have also separately evolved unique mechanisms to target CD45. While HCMV and MCMV target CD45 signaling and trafficking, HHV6A acts to downregulate CD45 transcripts. IMPORTANCE Human herpesviruses-6 and -7 infect essentially 100% of the world's population before the age of 5 and then remain latent or persistent in their host throughout life. As such, these viruses are among the most pervasive and stealthy of all viruses. Host immune cells rely on the presence of surface-expressed proteins to identify and target virus-infected cells. Here, we investigated the changes that occur to proteins expressed on the cell surface of T cells after infection with human herpesvirus-6A. We discovered that HHV-6A infection results in a reduction of CD45 on the surface of infected T cells and impaired activation in response to T cell receptor stimulation.


Assuntos
Regulação Enzimológica da Expressão Gênica , Regulação Viral da Expressão Gênica , Herpesvirus Humano 6/genética , Herpesvirus Humano 7/genética , Antígenos Comuns de Leucócito/genética , Linfócitos T/virologia , Linhagem Celular , Regulação para Baixo , Células HEK293 , Herpesvirus Humano 6/metabolismo , Herpesvirus Humano 7/metabolismo , Humanos , Estabilidade Proteica , Receptores de Antígenos de Linfócitos T/metabolismo , Linfócitos T/imunologia , Linfócitos T/metabolismo
12.
PLoS Comput Biol ; 17(5): e1007986, 2021 05.
Artigo em Inglês | MEDLINE | ID: mdl-34014917

RESUMO

The adaptive immune system serves as a potent and highly specific defense mechanism against pathogen infection. One component of this system, the effector T cell, facilitates pathogen clearance upon detection of specific antigens by the T cell receptor (TCR). A critical process in effector T cell activation is transmission of signals from the TCR to a key transcriptional regulator, NF-κB. The transmission of this signal involves a highly dynamic process in which helical filaments of Bcl10, a key protein constituent of the TCR signaling cascade, undergo competing processes of polymeric assembly and macroautophagy-dependent degradation. Through computational analysis of three-dimensional, super-resolution optical micrographs, we quantitatively characterize TCR-stimulated Bcl10 filament assembly and length dynamics, and demonstrate that filaments become shorter over time. Additionally, we develop an image-based, bootstrap-like resampling method that demonstrates the preferred association between autophagosomes and both Bcl10-filament ends and punctate-Bcl10 structures, implying that autophagosome-driven macroautophagy is directly responsible for Bcl10 filament shortening. We probe Bcl10 polymerization-depolymerization dynamics with a stochastic Monte-Carlo simulation of nucleation-limited filament assembly and degradation, and we show that high probabilities of filament nucleation in response to TCR engagement could provide the observed robust, homogeneous, and tunable response dynamic. Furthermore, we demonstrate that the speed of filament disassembly preferentially at filament ends provides effective regulatory control. Taken together, these data suggest that Bcl10 filament growth and degradation act as an excitable system that provides a digital response mechanism and the reliable timing critical for T cell activation and regulatory processes.


Assuntos
Proteína 10 de Linfoma CCL de Células B/metabolismo , Ativação Linfocitária , Linfócitos T/imunologia , Linfócitos T/metabolismo , Algoritmos , Animais , Autofagossomos/imunologia , Autofagossomos/metabolismo , Proteína 10 de Linfoma CCL de Células B/química , Proteína 10 de Linfoma CCL de Células B/genética , Linhagem Celular , Biologia Computacional , Simulação por Computador , Camundongos , Modelos Biológicos , Método de Monte Carlo , Polimerização , Proteólise , Receptores de Antígenos de Linfócitos T/metabolismo , Proteínas Recombinantes de Fusão/química , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismo , Transdução de Sinais
13.
Nat Immunol ; 22(6): 687-698, 2021 06.
Artigo em Inglês | MEDLINE | ID: mdl-33986548

RESUMO

The aged adaptive immune system is characterized by progressive dysfunction as well as increased autoimmunity. This decline is responsible for elevated susceptibility to infection and cancer, as well as decreased vaccination efficacy. Recent evidence indicates that CD4+ T cell-intrinsic alteratins contribute to chronic inflammation and are sufficient to accelerate an organism-wide aging phenotype, supporting the idea that T cell aging plays a major role in body-wide deterioration. In this Review, we propose ten molecular hallmarks to represent common denominators of T cell aging. These hallmarks are grouped into four primary hallmarks (thymic involution, mitochondrial dysfunction, genetic and epigenetic alterations, and loss of proteostasis) and four secondary hallmarks (reduction of the TCR repertoire, naive-memory imbalance, T cell senescence, and lack of effector plasticity), and together they explain the manifestation of the two integrative hallmarks (immunodeficiency and inflammaging). A major challenge now is weighing the relative impact of these hallmarks on T cell aging and understanding their interconnections, with the final goal of defining molecular targets for interventions in the aging process.


Assuntos
Envelhecimento/imunologia , Imunidade Celular , Linfócitos T/imunologia , Envelhecimento/genética , Autoimunidade/genética , Plasticidade Celular/genética , Plasticidade Celular/imunologia , Senescência Celular/genética , Senescência Celular/imunologia , Suscetibilidade a Doenças/imunologia , Epigênese Genética/imunologia , Regulação da Expressão Gênica/imunologia , Humanos , Inflamação/genética , Inflamação/imunologia , Proteostase/genética , Proteostase/imunologia , Receptores de Antígenos de Linfócitos T/genética , Receptores de Antígenos de Linfócitos T/metabolismo , Linfócitos T/metabolismo , Timo/imunologia , Timo/fisiopatologia
14.
Science ; 372(6543)2021 05 14.
Artigo em Inglês | MEDLINE | ID: mdl-33986151

RESUMO

Reactive oxygen species (ROS) increase in activated T cells because of metabolic activity induced to support T cell proliferation and differentiation. We show that these ROS trigger an oxidative stress response that leads to translation repression. This response is countered by Schlafen 2 (SLFN2), which directly binds transfer RNAs (tRNAs) to protect them from cleavage by the ribonuclease angiogenin. T cell-specific SLFN2 deficiency results in the accumulation of tRNA fragments, which inhibit translation and promote stress-granule formation. Interleukin-2 receptor ß (IL-2Rß) and IL-2Rγ fail to be translationally up-regulated after T cell receptor stimulation, rendering SLFN2-deficient T cells insensitive to interleukin-2's mitogenic effects. SLFN2 confers resistance against the ROS-mediated translation-inhibitory effects of oxidative stress normally induced by T cell activation, permitting the robust protein synthesis necessary for T cell expansion and immunity.


Assuntos
Proteínas de Ciclo Celular/metabolismo , Imunidade Celular , Estresse Oxidativo , RNA de Transferência/metabolismo , Linfócitos T/imunologia , Animais , Proteínas de Ciclo Celular/genética , Proliferação de Células , Feminino , Deleção de Genes , Infecções por Herpesviridae/imunologia , Subunidade gama Comum de Receptores de Interleucina/genética , Subunidade gama Comum de Receptores de Interleucina/metabolismo , Interleucina-2/metabolismo , Subunidade beta de Receptor de Interleucina-2/genética , Subunidade beta de Receptor de Interleucina-2/metabolismo , Ativação Linfocitária , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Muromegalovirus , Ligação Proteica , Biossíntese de Proteínas , Espécies Reativas de Oxigênio/metabolismo , Receptores de Antígenos de Linfócitos T/imunologia , Receptores de Antígenos de Linfócitos T/metabolismo , Ribonuclease Pancreático/genética , Ribonuclease Pancreático/metabolismo , Transdução de Sinais
15.
J Clin Immunol ; 41(6): 1131-1145, 2021 08.
Artigo em Inglês | MEDLINE | ID: mdl-33950324

RESUMO

The coronavirus disease 2019 (COVID-19) pandemic, caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has become a public health emergency. The most common symptoms of COVID-19 are fever, cough, and fatigue. While most patients with COVID-19 present with mild illness, some patients develop pneumonia, an important risk factor for mortality, at early stage of viral infection, putting these patients at increased risk of death. So far, little has been known about differences in the T cell repertoires between COVID-19 patients with and without pneumonia during SARS-CoV-2 infection. Herein, we aimed to investigate T cell receptor (TCR) repertoire profiles and patient-specific SARS-CoV-2-associated TCR clusters between COVID-19 patients with mild disease (no sign of pneumonia) and pneumonia. The TCR sequencing was conducted to characterize the peripheral TCR repertoire profile and diversity. The TCR clustering and CDR3 annotation were exploited to further discover groups of patient-specific TCR clonotypes with potential SARS-CoV-2 antigen specificities. Our study indicated a slight decrease in the TCR repertoire diversity and a skewed CDR3 length usage in patients with pneumonia compared to those with mild disease. The SARS-CoV-2-associated TCR clusters enriched in patients with mild disease exhibited significantly higher TCR generation probabilities and most of which were highly shared among patients, compared with those from pneumonia patients. Importantly, using similarity network-based clustering followed by the sequence conservation analysis, we found different patterns of CDR3 sequence motifs between mild disease- and pneumonia-specific SARS-CoV-2-associated public TCR clusters. Our results showed that characteristics of overall TCR repertoire and SARS-CoV-2-associated TCR clusters/clonotypes were divergent between COVID-19 patients with mild disease and patients with pneumonia. These findings provide important insights into the correlation between the TCR repertoire and disease severity in COVID-19 patients.


Assuntos
COVID-19/imunologia , Pneumonia/imunologia , Receptores de Antígenos de Linfócitos T/genética , SARS-CoV-2/fisiologia , Linfócitos T/imunologia , Adulto , Idoso , Progressão da Doença , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Pandemias , Receptores de Antígenos de Linfócitos T/metabolismo , Análise de Sequência de DNA , Índice de Gravidade de Doença
16.
Nucleic Acids Res ; 49(10): 5760-5778, 2021 06 04.
Artigo em Inglês | MEDLINE | ID: mdl-34037780

RESUMO

Alternative pre-mRNA splicing is a critical step to generate multiple transcripts, thereby dramatically enlarging the proteomic diversity. Thus, a common feature of most alternative splicing factor knockout models is lethality. However, little is known about lineage-specific alternative splicing regulators in a physiological setting. Here, we report that NSrp70 is selectively expressed in developing thymocytes, highest at the double-positive (DP) stage. Global splicing and transcriptional profiling revealed that NSrp70 regulates the cell cycle and survival of thymocytes by controlling the alternative processing of various RNA splicing factors, including the oncogenic splicing factor SRSF1. A conditional-knockout of Nsrp1 (NSrp70-cKO) using CD4Cre developed severe defects in T cell maturation to single-positive thymocytes, due to insufficient T cell receptor (TCR) signaling and uncontrolled cell growth and death. Mice displayed severe peripheral lymphopenia and could not optimally control tumor growth. This study establishes a model to address the function of lymphoid-lineage-specific alternative splicing factor NSrp70 in a thymic T cell developmental pathway.


Assuntos
Processamento Alternativo/genética , Carcinogênese/metabolismo , Desenvolvimento Embrionário/genética , Hematopoese/genética , Melanoma/metabolismo , Timócitos/metabolismo , Animais , Antígenos CD/metabolismo , Antígenos de Diferenciação de Linfócitos T/metabolismo , Apoptose/genética , Carcinogênese/genética , Proliferação de Células/genética , Genômica , Células HEK293 , Humanos , Lectinas Tipo C/metabolismo , Linfopenia/genética , Linfopenia/metabolismo , Melanoma/genética , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Reação em Cadeia da Polimerase , RNA-Seq , Reação em Cadeia da Polimerase em Tempo Real , Receptores de Antígenos de Linfócitos T/metabolismo , Fatores de Processamento de Serina-Arginina/genética , Fatores de Processamento de Serina-Arginina/metabolismo , Timo/embriologia , Timo/metabolismo
17.
Nat Commun ; 12(1): 2746, 2021 05 12.
Artigo em Inglês | MEDLINE | ID: mdl-33980853

RESUMO

Despite the clinical success of blocking its interactions, how PD-1 inhibits T-cell activation is incompletely understood, as exemplified by its potency far exceeding what might be predicted from its affinity for PD-1 ligand-1 (PD-L1). This may be partially attributed to PD-1's targeting the proximal signaling of the T-cell receptor (TCR) and co-stimulatory receptor CD28 via activating Src homology region 2 domain-containing phosphatases (SHPs). Here, we report PD-1 signaling regulates the initial TCR antigen recognition manifested in a smaller spreading area, fewer molecular bonds formed, and shorter bond lifetime of T cell interaction with peptide-major histocompatibility complex (pMHC) in the presence than absence of PD-L1 in a manner dependent on SHPs and Leukocyte C-terminal Src kinase. Our results identify a PD-1 inhibitory mechanism that disrupts the cooperative TCR-pMHC-CD8 trimolecular interaction, which prevents CD8 from augmenting antigen recognition, explaining PD-1's potent inhibitory function and its value as a target for clinical intervention.


Assuntos
Antígenos CD8/imunologia , Receptor de Morte Celular Programada 1/imunologia , Receptores de Antígenos de Linfócitos T/imunologia , Linfócitos T/imunologia , Animais , Antígeno B7-H1/imunologia , Antígenos CD8/metabolismo , Cálcio/metabolismo , Humanos , Ativação Linfocitária , Proteína Tirosina Quinase p56(lck) Linfócito-Específica/metabolismo , Complexo Principal de Histocompatibilidade/imunologia , Camundongos , Camundongos Transgênicos , Receptor de Morte Celular Programada 1/genética , Receptor de Morte Celular Programada 1/metabolismo , Ligação Proteica , Receptores de Antígenos de Linfócitos T/metabolismo , Transdução de Sinais , Linfócitos T/metabolismo
19.
J Immunol ; 206(9): 2045-2051, 2021 05 01.
Artigo em Inglês | MEDLINE | ID: mdl-33846228

RESUMO

Autoreactive CD4 T cells are thought to play pivotal roles in the pathogenesis of rheumatoid arthritis (RA). Recently, a subset of CD4 T cells that express high levels of programmed death-1 (PD-1) but are distinct from follicular helper T cells have been identified in the joints of RA patients and named peripheral helper T (Tph) cells. Because PD-1 is expressed on T cells chronically stimulated with the Ags, we tested a hypothesis that Tph cells are the pathogenic autoreactive CD4 T cells in RA. We found that human Tph cells in RA joints produce proinflammatory effector cytokines, including IFN-γ, TNF-α, and GM-CSF, in addition to B cell-helping cytokines, such as IL-21 and CXCL13. Flow cytometric analysis showed different bias of TCR Vß usage between PD-1high Tph cells and PD-1low/neg CD4 T cells, including Th1 cells, in the joint or memory CD4 T cells in the peripheral blood, whereas there was little difference between the latter two subsets. In line with this, deep sequencing of TCR demonstrated an overlap of expanded clones between peripheral blood memory CD4 T cells and PD-1low/neg CD4 T cells but not Tph cells in the joint. Interestingly, Tph cells preferentially exhibited autologous MLR in vitro, which required recognition of self-MHC class II and was pronounced by blocking PD-1 signaling. Taken together, these results suggest that Tph cells are the pathogenic autoreactive CD4 T cells in RA, which expand locally in the joints and are regulated by PD-1 signaling.


Assuntos
Artrite Reumatoide/imunologia , Linfócitos T CD4-Positivos/imunologia , Receptor de Morte Celular Programada 1/imunologia , Linfócitos T Auxiliares-Indutores/imunologia , Idoso , Artrite Reumatoide/metabolismo , Linfócitos T CD4-Positivos/metabolismo , Células Cultivadas , Quimiocina CXCL13/imunologia , Quimiocina CXCL13/metabolismo , Citocinas/imunologia , Citocinas/metabolismo , Feminino , Humanos , Mediadores da Inflamação/imunologia , Mediadores da Inflamação/metabolismo , Masculino , Pessoa de Meia-Idade , Receptor de Morte Celular Programada 1/metabolismo , Receptores de Antígenos de Linfócitos T/imunologia , Receptores de Antígenos de Linfócitos T/metabolismo , Transdução de Sinais/imunologia , Linfócitos T Auxiliares-Indutores/metabolismo , Células Th1/imunologia , Células Th1/metabolismo
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