The peculiarity of T cell is their ability to recognize an infinite range of self and foreign antigens. This ability is achieved during thymic development through a complex molecular mechanism based ...on somatic recombination that leads to the expression of a very heterogeneous population of surface antigen receptors, the T Cell Receptors (TCRs). TCRs are cell specific and represent a sort of "molecular tag" of T cells and have been widely studied to monitor the dynamics of T cells in terms of clonality and diversity in several contexts including lymphoid malignancies, infectious diseases, autoimmune diseases, and tumor immunology. In this review, we provide an overview of the strategies used to investigate the TCR repertoire from the pioneering techniques based on the V segments identification to the revolution introduced by Next-Generation Sequencing that allows for high-throughput sequencing of alpha and beta chains. Single cell based approaches brought the analysis to a higher level of complexity and now provide the opportunity to sequence paired alpha and beta chains. We also discuss novel approaches that through the integration of TCR tracking and mRNA single cell sequencing offer a valuable tool to associate antigen specificity to transcriptional dynamics and to understand the molecular mechanisms of T cell plasticity.
Regulatory T cells (Tregs) are critical for maintaining immune homeostasis, but their presence in tumor tissues impairs anti-tumor immunity and portends poor prognoses in cancer patients. Here, we ...reveal a mechanism to selectively target and reprogram the function of tumor-infiltrating Tregs (TI-Tregs) by exploiting their dependency on the histone H3K27 methyltransferase enhancer of zeste homolog 2 (EZH2) in tumors. Disruption of EZH2 activity in Tregs, either pharmacologically or genetically, drove the acquisition of pro-inflammatory functions in TI-Tregs, remodeling the tumor microenvironment and enhancing the recruitment and function of CD8+ and CD4+ effector T cells that eliminate tumors. Moreover, abolishing EZH2 function in Tregs was mechanistically distinct from, more potent than, and less toxic than a generalized Treg depletion approach. This study reveals a strategy to target Tregs in cancer that mitigates autoimmunity by reprogramming their function in tumors to enhance anti-cancer immunity.
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•EZH2 expression is elevated in tumor-infiltrating (TI) Tregs•Pharmacological inhibition of EZH2 destabilizes FOXP3 expression and slows tumor growth•Genetic disruption of Ezh2 function in Tregs leads to robust anti-tumor immunity•Blockade of EZH2 in Tregs reprograms TI-Tregs to gain pro-inflammatory activity
EZH2 plays an intrinsic role in neoplastic cells as an oncogene, prompting the development of EZH2 inhibitors for cancer therapy. Wang et al. show that disrupting EZH2 function also has immunomodulatory activities and, when blocked in Tregs, promotes potent cancer immunity.
The molecular mechanisms responsible for the high immunosuppressive capacity of CD4+ Tregs in tumors are not well known. High-dimensional single-cell profiling of T cells from chemotherapy-naive ...individuals with non-small-cell lung cancer identified the transcription factor IRF4 as specifically expressed by a subset of intratumoral CD4+ effector Tregs with superior suppressive activity. In contrast to the IRF4- counterparts, IRF4+ Tregs expressed a vast array of suppressive molecules, and their presence correlated with multiple exhausted subpopulations of T cells. Integration of transcriptomic and epigenomic data revealed that IRF4, either alone or in combination with its partner BATF, directly controlled a molecular program responsible for immunosuppression in tumors. Accordingly, deletion of Irf4 exclusively in Tregs resulted in delayed tumor growth in mice while the abundance of IRF4+ Tregs correlated with poor prognosis in patients with multiple human cancers. Thus, a common mechanism underlies immunosuppression in the tumor microenvironment irrespective of the tumor type.
Multiple sclerosis (MS) is a T cell driven autoimmune disease of the central nervous system (CNS). Despite its association with Epstein-Barr Virus (EBV), how viral infections promote MS remains ...unclear. However, there is increasing evidence that the CNS is continuously surveyed by virus-specific T cells, which protect against reactivating neurotropic viruses. Here, we discuss how viral infections could lead to the breakdown of self-tolerance in genetically predisposed individuals, and how the reactivations of viruses in the CNS could induce the recruitment of both autoaggressive and virus-specific T cell subsets, causing relapses and progressive disability. A disturbed immune surveillance in MS would explain several experimental findings, and has important implications for prognosis and therapy.
A huge body of evidence suggests that viral infections promote MS; however, no single causal virus has been identified. Multiple viruses could promote MS via bystander effects.
Molecular mimicry is an established pathogenic mechanism in selected autoimmune diseases. It is also well documented in MS, but its contribution to MS pathogenesis is still unclear.
Bystander activation upon viral infection could be involved in the generation of the autoreactive and potentially encephalitogenic T helper (Th)-1/17 central memory (Th1/17CM) cells found in the circulation of patients with MS.
Autoreactive Th1/17CM cells could expand at the cost of antiviral Th1CM cells in patients with MS, in particular in those undergoing natalizumab therapy, because these cells are expected to compete for the same homeostatic niche.
Autoreactive Th1/17 cells and antiviral Th1 cells are recruited to the CSF of patients with MS following attacks, suggesting that viral reactivations in the CNS induce the recruitment of pathogenic Th1/17 cells. Autoreactive Th1/17 cells in the CNS might also induce de novo viral reactivations in a circuit of self-induced inflammation.
The aim was to investigate CD4
T-cell subsets, immune cells and their cytokine profiles in blood and synovial compartments in rheumatoid arthritis (RA) and inflammatory osteoarthritis (OA) to define ...specific immune signatures.
Peripheral blood, synovial fluid (SF) and synovial membranes (SM) of RA and OA patients were analyzed. CD4
T-cell subset frequencies were determined by flow cytometry, and cytokine concentrations in serum and SF were measured by ELISA.
In peripheral blood, OA patients had altered frequencies of regulatory T-cell subsets, and higher frequencies of Th17 and of Th1/17 cells than RA patients. In the synovial compartment of OA patients, conventional Th17 cells were largely excluded, while Th1/17 cells were enriched and more frequent than in RA patients. Conversely, in the synovial compartment of RA patients, regulatory T cells and Tfh cells were enriched and more frequent then in OA patients. IL-17 and Blys were increased both in serum and SF of RA patients, and correlated with autoantibodies and disease activity. Notably, Blys levels were already significantly elevated in RA patients with low disease activity score in 28 joints (DAS28) and without autoantibody positivity.
Although patients with inflammatory OA have immune activation in the synovial compartment, they display different T-cell subset frequencies and cytokine profiles. Soluble mediators such as Blys might help to discriminate mild clinical forms of RA from inflammatory OA particularly at the onset of the disease.
Highlights • IL-10 produced by regulatory T-cells has a non-redundant role to maintain intestinal immune homeostasis and to prevent colitis. • IL-10 produced by B helper T-cells promotes autoantibody ...production, and has a pathogenic role in systemic lupus erythematosus. • IL-10 produced early by regulatory T-cells or late by helper T-cells has opposing functions in cytotoxic T-cell responses and cancer.
Regulatory T (T
) cells are a barrier for tumor immunity and a target for immunotherapy. Using single-cell transcriptomics, we found that CD4
T cells infiltrating primary and metastatic colorectal ...cancer and non-small-cell lung cancer are highly enriched for two subsets of comparable size and suppressor function comprising forkhead box protein P3
T
and eomesodermin homolog (EOMES)
type 1 regulatory T (Tr1)-like cells also expressing granzyme K and chitinase-3-like protein 2. EOMES
Tr1-like cells, but not T
cells, were clonally related to effector T cells and were clonally expanded in primary and metastatic tumors, which is consistent with their proliferation and differentiation in situ. Using chitinase-3-like protein 2 as a subset signature, we found that the EOMES
Tr1-like subset correlates with disease progression but is also associated with response to programmed cell death protein 1-targeted immunotherapy. Collectively, these findings highlight the heterogeneity of T
cells that accumulate in primary tumors and metastases and identify a new prospective target for cancer immunotherapy.
Deciphering how the human striatum develops is necessary for understanding the diseases that affect this region. To decode the transcriptional modules that regulate this structure during development, ...we compiled a catalog of 1116 long intergenic noncoding RNAs (lincRNAs) identified de novo and then profiled 96,789 single cells from the early human fetal striatum. We found that D1 and D2 medium spiny neurons (D1- and D2-MSNs) arise from a common progenitor and that lineage commitment is established during the postmitotic transition, across a pre-MSN phase that exhibits a continuous spectrum of fate determinants. We then uncovered cell type-specific gene regulatory networks that we validated through in silico perturbation. Finally, we identified human-specific lincRNAs that contribute to the phylogenetic divergence of this structure in humans. This work delineates the cellular hierarchies governing MSN lineage commitment.
4-anilino quinazolines have been identified as inhibitors of HCV replication. The target of this class of compounds was proposed to be the viral protein NS5A, although unequivocal proof has never ...been presented. A 4-anilino quinazoline moiety is often found in kinase inhibitors, leading us to formulate the hypothesis that the anti-HCV activity displayed by these compounds might be due to inhibition of a cellular kinase. Type III phosphatidylinositol 4-kinase α (PI4KIIIα) has recently been identified as a host factor for HCV replication. We therefore evaluated AL-9, a compound prototypical of the 4-anilino quinazoline class, on selected phosphatidylinositol kinases. AL-9 inhibited purified PI4KIIIα and, to a lesser extent, PI4KIIIβ. In Huh7.5 cells, PI4KIIIα is responsible for the phosphatidylinositol-4 phosphate (PI4P) pool present in the plasma membrane. Accordingly, we observed a gradual decrease of PI4P in the plasma membrane upon incubation with AL-9, indicating that this agent inhibits PI4KIIIα also in living cells. Conversely, AL-9 did not affect the level of PI4P in the Golgi membrane, suggesting that the PI4KIIIβ isoform was not significantly inhibited under our experimental conditions. Incubation of cells expressing HCV proteins with AL-9 induced abnormally large clusters of NS5A, a phenomenon previously observed upon silencing PI4KIIIα by RNA interference. In light of our findings, we propose that the antiviral effect of 4-anilino quinazoline compounds is mediated by the inhibition of PI4KIIIα and the consequent depletion of PI4P required for the HCV membranous web. In addition, we noted that HCV has a profound effect on cellular PI4P distribution, causing significant enrichment of PI4P in the HCV-membranous web and a concomitant depletion of PI4P in the plasma membrane. This observation implies that HCV--by recruiting PI4KIIIα in the RNA replication complex--hijacks PI4P metabolism, ultimately resulting in a markedly altered subcellular distribution of the PI4KIIIα product.
The basic unit of genome packaging is the nucleosome, and nucleosomes have long been proposed to restrict DNA accessibility both to damage and to transcription. Nucleosome number in cells was ...considered fixed, but recently aging yeast and mammalian cells were shown to contain fewer nucleosomes. We show here that mammalian cells lacking High Mobility Group Box 1 protein (HMGB1) contain a reduced amount of core, linker, and variant histones, and a correspondingly reduced number of nucleosomes, possibly because HMGB1 facilitates nucleosome assembly. Yeast nhp6 mutants lacking Nhp6a and -b proteins, which are related to HMGB1, also have a reduced amount of histones and fewer nucleosomes. Nucleosome limitation in both mammalian and yeast cells increases the sensitivity of DNA to damage, increases transcription globally, and affects the relative expression of about 10% of genes. In yeast nhp6 cells the loss of more than one nucleosome in four does not affect the location of nucleosomes and their spacing, but nucleosomal occupancy. The decrease in nucleosomal occupancy is non-uniform and can be modelled assuming that different nucleosomal sites compete for available histones. Sites with a high propensity to occupation are almost always packaged into nucleosomes both in wild type and nucleosome-depleted cells; nucleosomes on sites with low propensity to occupation are disproportionately lost in nucleosome-depleted cells. We suggest that variation in nucleosome number, by affecting nucleosomal occupancy both genomewide and gene-specifically, constitutes a novel layer of epigenetic regulation.
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