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The transcriptional and epigenetic regulation of CD8+ T cell differentiation is critical for balancing pathogen eradication and long-term immunity by effector and memory CTLs, respectively. In this study, we demonstrate that the lysine demethylase 6b (Kdm6b) is essential for the proper generation and function of effector CD81 T cells during acute infection and tumor eradication. We found that cells lacking Kdm6b (by either T cell-specific knockout mice or knockdown using short hairpin RNA strategies) show an enhanced generation of memory precursor and early effector cells upon acute viral infection in a cell-intrinsic manner. We also demonstrate that Kdm6b is indispensable for proper effector functions and tumor protection, and that memory CD8+ T cells lacking Kdm6b displayed a defective recall response. Mechanistically, we identified that Kdm6b, through induction of chromatin accessibility in key effector-associated gene loci, allows for the proper generation of effector CTLs. Our results pinpoint the essential function of Kdm6b in allowing chromatin accessibility in effector-associated genes, and identify Kdm6b as a potential target for therapeutics in diseases with dysregulated effector responses. [ABSTRACT FROM AUTHOR]

Protein phosphatase 2A (PP2A) composed of a scaffold subunit, a catalytic subunit, and multiple regulatory subunits is a ubiquitously expressed serine/threonine phosphatase. We have previously shown that the PP2A catalytic subunit is increased in T cells from patients with systemic lupus erythematosus and promotes IL-17 production by enhancing the activity of Rho-associated kinase (ROCK) in T cells. However, the molecular mechanism whereby PP2A regulates ROCK activity is unknown. In this study, we show that the PP2A regulatory subunit PPP2R2A is increased in T cells from people with systemic lupus erythematosus and binds to, dephosphorylates, and activates the guanine nucleotide exchange factor GEF-H1 at Ser885, which in turn increases the levels of RhoA-GTP and the activity of ROCK in T cells. Genetic PPP2R2A deficiency in murine T cells reduced Th1 and Th17, but not regulatory T cell differentiation and mice with T cell-specific PPP2R2A deficiency displayed less autoimmunity when immunized with myelin oligodendrocyte glycoprotein peptide. Our studies indicate that PPP2R2A is the regulatory subunit that dictates the PP2A-directed enhanced Th1 and Th17 differentiation, and therefore, it represents a therapeutic target for pathologies linked to Th1 and Th17 cell expansion. [ABSTRACT FROM AUTHOR]

Foxo1 is an essential transcription factor required for the survival and differentiation of memory CD8 T cells, yet it is unclear whether these Foxo1-dependent functions are inherently coupled. To address this question, we examined the effects of different Foxo1 posttranslational modifications. Phosphorylation of Foxo1 by Akt kinases at three distinct residues is well characterized to inhibit Foxo1 transcriptional activity. However, the effect of Foxo1 phosphorylation within its DNA-binding domain at serine 209 by Mst1 kinase is not fully understood. In this study, we show that an S209A phospho-null Foxo1 exhibited Akt-dependent nuclear trafficking in mouse CD8 T cells and augmented the expression of canonical Foxo1 target genes such as Il7r and Sell. In contrast, an S209D phosphomimetic Foxo1 (SD-Foxo1) was largely excluded from the nucleus of CD8 T cells and failed to transactivate these genes. RNA sequencing analysis revealed that SD-Foxo1 was associated with a distinct Foxo1-dependent transcriptional profile, including genes mediating CD8 effector function and cell survival. Despite defective transactivation of canonical target genes, SD-Foxo1 promoted IL-15-mediated CD8 T cell survival in vitro and survival of short-lived effector cells in vivo in response to Listeria monocytogenes infection. However, SD-Foxo1 actively repressed CD127 expression and failed to generate memory precursors and long-lived memory T cells. Together, these data indicate that S209 is a critical residue for the regulation of Foxo1 subcellular localization and for balancing CD8 T cell differentiation and survival. [ABSTRACT FROM AUTHOR]

Immune homeostasis in peripheral tissues is, to a large degree, maintained by the differentiation and action of regulatory T cells (Treg) specific for tissue Ags. Using a novel mouse model, we have studied the differentiation of naive CD4+ T cells into Foxp3+ Treg in response to a cutaneous Ag (OVA). We found that expression of OVA resulted in fatal autoimmunity and in prevention of peripheral Treg generation. Inhibiting mTOR activity with rapamycin rescued the generation of Foxp3+ T cells. When we varied the level of Ag expression to modulate TCR signaling, we found that low Ag concentrations promoted the generation of Foxp3+ T cells, whereas high levels expanded effector T cells and caused severe autoimmunity. Our findings indicate that the expression level of tissue Ag is a key determinant of the balance between tissue-reactive effector and peripheral Foxp3+ T cells, which determines the choice between tolerance and autoimmunity. [ABSTRACT FROM AUTHOR]

CD8 T cell differentiation is orchestrated by dynamic metabolic changes that direct activation, proliferation, cytotoxic function, and epigenetic changes. We report that the BTB-ZF family transcriptional repressor Zbtb20 negatively regulates CD8 T cell metabolism and memory differentiation in mice. Effector and memory CD8 T cells with conditional Zbtb20 deficiency displayed enhanced mitochondrial and glycolytic metabolism, and memory CD8 T cells had enhanced spare respiratory capacity. Furthermore, Zbtb20-deficient CD8 T cells displayed increased flexibility in the use of mitochondrial fuel sources. Phenotypic and transcriptional skewing toward the memory fate was observed during the CD8 T cell response to Listeria monocytogenes. Memory cells mounted larger secondary responses and conferred better protection following tumor challenge. These data suggest that inactivation of Zbtb20 may offer an approach to enhance metabolic activity and flexibility and improve memory CD8 T cell differentiation, useful attributes for T cells used in adoptive immunotherapy. [ABSTRACT FROM AUTHOR]

The roles distinct B cell subsets play in clonal expansion, isotype switching, and memory B cell differentiation in response to T cell-independent type 2 Ags (TI-2 Ags) has been understudied. Using sorted B cells from VHB1-8 knock-in mice, we evaluated B-1b, marginal zone, and follicular B cell responses to the TI-2 Ag, NP-Ficoll. All subsets extensively divided in response to NP-Ficoll. Nonetheless, B-1b cells exhibited significantly increased IgG switching and differentiation into Ab-secreting cells (ASC)--a finding that coincided with increased AgR signaling capacity and Blimp1 expression by B-1b cells. All subsets formed memory cells and expressed markers previously identified for T cell-dependent memory B cells, including CD80, PDL2, and CD73, although B-1b cells generated the greatest number of memory cells with higher frequencies of IgG- and CD80-expressing cells. Despite memory formation, secondary immunization 4 wk after primary immunization did not increase NP-specific IgG. However, boosting occurred in B-1b cell-recipient mice when IgG levels declined. CD80+ memory B-1b cells divided, class switched, and differentiated into ASC in response to Ag in vivo, but this was inhibited in the presence of NP-specific IgG. Furthermore, CD80 blockade significantly increased memory B-1b cell division and differentiation to ASC upon Ag restimulation. Collectively, these findings demonstrate B-1b, marginal zone B, and follicular B subsets significantly contribute to the TI-2 Ag-specific memory B cell pool. In particular, we show B-1b cells generate a functional CD80-regulated memory population that can be stimulated to divide and differentiate into ASC upon Ag re-encounter when Ag-specific IgG levels decline. [ABSTRACT FROM AUTHOR]

Despite the fact that the majority of people in tuberculosis (TB)-endemic areas are vaccinated with the Bacillus Calmette-Guérin (BCG) vaccine, TB remains the leading infectious cause of death. Data from both animal models and humans show that BCG and subunit vaccines induce T cells of different phenotypes, and little is known about how BCG priming influences subsequent booster vaccines. To test this, we designed a novel Mycobacterium tuberculosis-specific (or "non-BCG") subunit vaccine with protective efficacy in both mice and guinea pigs and compared it to a known BCG boosting vaccine. In naive mice, this M. tuberculosis-specific vaccine induced similar protection compared with the BCG boosting vaccine. However, in BCG-primed animals, only the M. tuberculosis-specific vaccine added significantly to the BCG-induced protection. This correlated with the priming of T cells with a lower degree of differentiation and improved lung-homing capacity. These results have implications for TB vaccine design. [ABSTRACT FROM AUTHOR]

CD8+ T cells are critical mediators of adaptive immunity, and enhancing their function can promote robust responses against invading pathogens and neoplastic cells. In addition to TCR stimulation, the provision of costimulation through ligation of TNFR family members, such as OX40 (CD134), provides essential signals driving T cell differentiation, survival, and memory in part through enhanced IL-2/IL-2R signaling. Interestingly, TCR stimulation in the presence of IL-2 upregulates intracellular expression of the b-galactoside binding protein, Galectin-3 (Gal-3). Gal-3 has been shown to regulate Th1/Th2 polarization of CD4+ T cells; however, the extent to which Gal-3 regulates the OX40/IL-2 signaling axis and CD8+ T cell proliferation, effector function, and/or survival is unknown. In this study, we demonstrate that murine Gal-3-deficient CD8+ T cells exhibited no defects in early (36 h) activation or proliferation following TCR stimulation. In contrast, Gal-32/2 CD8+ T cells exhibited decreased survival and a reduced capacity to develop into memory cells following stimulation with cognate Ag plus agonist anti-OX40 mAb or IL-2 in vivo. Decreased survival of Gal-3-/-T cells was associated with increased apoptosis and occurred in a cell-intrinsic manner. Together, these data implicate intracellular Gal-3 as a critical mediator of OX40-mediated CD8+ T cell survival and memory formation following Ag exposure. [ABSTRACT FROM AUTHOR]

Long noncoding RNAs (lncRNAs) have been demonstrated to play important regulatory roles in gene expression, from histone modification to protein stability. However, the functions of most identified lncRNAs are not known. In this study, we investigated the role of an lncRNA called AW112010. The expression of AW112010 was significantly increased in CD4+ T cells from C57BL/6J mice activated in vivo with myelin oligodendrocyte glycoprotein, Staphylococcal enterotoxin B, or in vitro with anti-CD3 anti-CD28 mAbs, thereby demonstrating that activation of T cells leads to induction of AW112010. In contrast, anti-inflammatory cannabinoids such as cannabidiol or d-9-tetrahydrocannabinol decreased the expression of AW112010 in T cells. Interestingly, the expression of AW112010 was high in in vitro-polarized Th1 and Th17 cells but low in Th2 cells, suggesting that this lncRNA may regulate inflammation. To identify genes that might be regulated by AW112010, we used chromatin isolation by RNA purification, followed by sequencing. This approach demonstrated that AW112010 regulated the transcription of IL-10. Additionally, the level of IL-10 in activated T cells was low when the expression of AW112010 was increased. Use of small interfering RNA to knock down AW112010 expression in activated T cells led to increased IL-10 expression and a decrease in the expression of IFN-g. Further studies showed that AW112010 interacted with histone demethylase KDM5A, which led to decreased H3K4 methylation in IL-10 gene locus. Together, these studies demonstrate that lncRNA AW112010 promotes the differentiation of inflammatory T cells by suppressing IL-10 expression through histone demethylation. [ABSTRACT FROM AUTHOR]

Common IRF5 genetic risk variants associated with multiple immune-mediated diseases are a major determinant of interindividual variability in pattern-recognition receptor (PRR)-induced cytokines in myeloid cells. However, how myeloid cell-intrinsic IRF5 regulates the multiple distinct checkpoints mediating T cell outcomes in vivo and IRF5-dependent mechanisms contributing to these distinct checkpoints are not well defined. Using an in vivo Ag-specific adoptive T cell transfer approach into Irf52/2 mice, we found that T cell-extrinsic IRF5 regulated T cell outcomes at multiple critical checkpoints, including chemokine-mediated T cell trafficking into lymph nodes and PDK1-dependent soluble Ag uptake, costimulatory molecule upregulation, and secretion of Th1 (IL-12)- and Th17 (IL-23, IL-1b, and IL-6)-conditioning cytokines by myeloid cells, which then cross-regulated Th2 and regulatory T cells. IRF5 was required for PRR-induced MAPK and NF-kB activation, which, in turn, regulated these key outcomes in myeloid cells. Importantly, mice with IRF5 deleted from myeloid cells demonstrated T cell outcomes similar to those observed in Irf52/2 mice. Complementation of IL-12 and IL-23 was able to restore T cell differentiation both in vitro and in vivo in the context of myeloid cell-deficient IRF5. Finally, human monocyte-derived dendritic cells from IRF5 disease-associated genetic risk carriers leading to increased IRF5 expression demonstrated increased Ag uptake and increased PRR-induced costimulatory molecule expression and chemokine and cytokine secretion compared with monocyte-derived dendritic cells from low-expressing IRF5 genetic variant carriers. These data establish that myeloid cell-intrinsic IRF5 regulates multiple distinct checkpoints in T cell activation and differentiation and that these are modulated by IRF5 disease risk variants. [ABSTRACT FROM AUTHOR]

Metabolic reprogramming plays a central role in T cell activation and differentiation, and the inhibition of key metabolic pathways in activated T cells represents a logical approach for the development of new therapeutic agents for treating autoimmune diseases. The widely prescribed antidiabetic drug metformin and the glycolytic inhibitor 2-deoxyglucose (2-DG) have been used to study the inhibition of oxidative phosphorylation and glycolysis, respectively, in murine immune cells. Published studies have demonstrated that combination treatment with metformin and 2-DG was efficacious in dampening mouse T cell activation-induced effector processes, relative to treatments with either metformin or 2-DG alone. In this study, we report that metformin + 2-DG treatment more potently suppressed IFN-g production and cell proliferation in activated primary human CD4+ T cells than either metformin or 2-DG treatment alone. The effects of metformin + 2-DG on human T cells were accompanied by significant remodeling of activation-induced metabolic transcriptional programs, in part because of suppression of key transcriptional regulators MYC and HIF-1A. Accordingly, metformin + 2-DG treatment significantly suppressed MYC-dependent metabolic genes and processes, but this effect was found to be independent of mTORC1 signaling. These findings reveal significant insights into the effects of metabolic inhibition by metformin + 2-DG treatment on primary human T cells and provide a basis for future work aimed at developing new combination therapy regimens that target multiple pathways within the metabolic networks of activated human T cells. [ABSTRACT FROM AUTHOR]

Metformin has been studied for its anticancer effects by regulating T cell functions. However, the mechanisms through which metformin stimulates the differentiation of memory T cells remain unclear. We found that the frequencies of memory stem and central memory T cells increased for both in peripheral and tumor-infiltrating CD8+ T cells in metformin-treated lung cancer patients compared with those not taking the medication. An in vitro assay showed that metformin promoted the formation of memory CD8+ T cells and enhanced their antiapoptotic abilities. In addition, AMP-activated protein kinase (AMPK) activation decreased microRNA-107 expression, thus enhancing Eomesodermin expression, which suppressed the transcription of PDCD1 in metformin-treated CD8+ T cells. In the CAR-T cell therapy model, metformin also exhibited cytotoxicity-promoting effects that led to decreased tumor growth. Metformin could reprogram the differentiation of CD8+ T cells, which may benefit the clinical therapy of cancer patients by facilitating long-lasting cytotoxic functions. [ABSTRACT FROM AUTHOR]

Type I IFN is produced upon infection and tissue damage and induces the expression of many IFN-stimulated genes (ISGs) that encode host-protective proteins. ISG15 is a ubiqitin-like molecule that can be conjugated to proteins but is also released from cells in a free form. Free, extracellular ISG15 is suggested to have an immune-regulatory role, based on disease phenotypes of ISG15-deficient humans and mice. However, the underlying mechanisms by which free ISG15 would act as a "cytokine" are unclear and much debated. We, in this study, demonstrate in a clinically relevant mouse model of therapeutic vaccination that free ISG15 is an alarmin that induces tissue alert, characterized by extracellular matrix remodeling, myeloid cell infiltration, and inflammation. Moreover, free ISG15 is a potent adjuvant for the CTL response. ISG15 produced at the vaccination site promoted the vaccine-specific CTL response by enhancing expansion, short-lived effector and effector/memory differentiation of CD8+ T cells. The function of free ISG15 as an extracellular ligand was demonstrated, because the equivalents in murine ISG15 of 2 aa recently implicated in binding of human ISG15 to LFA-1 in vitro were required for its adjuvant effect in vivo. Moreover, in further agreement with the in vitro findings on human cells, free ISG15 boosted the CTL response in vivo via NK cells in the absence of CD4+ T cell help. Thus, free ISG15 is part of a newly recognized innate route to promote the CTL response. [ABSTRACT FROM AUTHOR]

T cell-mediated immune response plays a crucial role in controlling Trypanosoma cruzi infection and parasite burden, but it is also involved in the clinical onset and progression of chronic Chagas' disease. Therefore, the study of T cells is central to the understanding of the immune response against the parasite and its implications for the infected organism. The complexity of the parasite-host interactions hampers the identification and characterization of T cell-activating epitopes. We approached this issue by combining in silico and in vitro methods to interrogate patients' T cells specificity. Fifty T. cruzi peptides predicted to bind a broad range of class I and II HLA molecules were selected for in vitro screening against PBMC samples from a cohort of chronic Chagas' disease patients, using IFN-γ secretion as a readout. Seven of these peptides were shown to activate this type of T cell response, and four out of these contain class I and II epitopes that, to our knowledge, are first described in this study. The remaining three contain sequences that had been previously demonstrated to induce CD8+ T cell response in Chagas' disease patients, or bind HLA-A*02:01, but are, in this study, demonstrated to engage CD4+ T cells. We also assessed the degree of differentiation of activated T cells and looked into the HLA variants that might restrict the recognition of these peptides in the context of human T. cruzi infection. [ABSTRACT FROM AUTHOR]

TCR signaling activates kinases including AKT/mTOR that engage metabolic networks to support the energetic demands of a T cell during an immune response. It is realized that CD4+ T cell subsets have different metabolic requirements. Yet, how TCR signaling is coupled to the regulation of intermediate metabolites and how changes in metabolite flux contribute to T cell differentiation are less established. We find that TCR signaling regulates acetyl-CoA metabolism via AKT in murine CD4+ T cells. Weak TCR signals promote AKT-catalyzed phosphorylation and inhibition of citrate synthase, elevated acetyl-CoA levels, and hyperacetylation of mitochondrial proteins. Genetic knockdown of citrate synthase promotes increased nuclear acetyl- CoA levels, increased histone acetylation at the FOXP3 promotor and induction of FOXP3 transcription. These data identify a circuit between AKT signaling and acetyl-CoA metabolism regulated via TCR signal strength and that transient fluctuations in acetyl-CoA levels function in T cell fate decisions. [ABSTRACT FROM AUTHOR]

Foxp3+ regulatory T (TREG) cells are central mediators in the control of peripheral immune responses. Genome-wide transcriptional profiles show canonical signatures for Foxp3+ TREG cells, distinguishing them from Foxp32 effector T (TEFF) cells. We previously uncovered distinct mRNA translational signatures differentiating CD4+ TEFF and TREG cells through parallel measurements of cytosolic (global) and polysome-associated (translationally enhanced) mRNA levels in both subsets. We show that the mRNA encoding for the ubiquitin-specific peptidase 11 (USP11), a known modulator of TGF-γ signaling, was preferentially translated in TCR-activated TREG cells compared with conventional, murine CD4+ T cells. TGF-b is a key cytokine driving the induction and maintenance of Foxp3 expression in T cells. We hypothesized that differential translation of USP11 mRNA endows TREG cells with an advantage to respond to TGF-b signals. In an in vivo mouse model promoting TREG cells plasticity, we found that USP11 protein was expressed at elevated levels in stable TREG cells, whereas ectopic USP11 expression enhanced the suppressive capacity and lineage commitment of these cells in vitro and in vivo. USP11 overexpression in TEFF cells enhanced the activation of the TGF-γ pathway and promoted TREG or TH17, but not Th1, cell differentiation in vitro and in vivo, an effect abrogated by USP11 gene silencing or the inhibition of enzymatic activity. Thus, USP11 potentiates TGF-b signaling in both TREG and TEFF cells, in turn driving increased suppressive function and lineage commitment in thymic-derived TREG cells and potentiating the TGF-b-dependent differentiation of TEFF cells to peripherally induced TREG and TH17 cells. [ABSTRACT FROM AUTHOR]

CMV remains an important opportunistic pathogen in high-risk lung transplant recipients. We characterized the phenotype and function of CD8+ T cells from acute/primary into chronic CMV infection in 23 (donor+/recipient2; D+R2) lung transplant recipients and found rapid induction of both KLRG1+ and/or CD57+ CMV-specific CD8+ T cells with unexpected coexpression of CD27. These cells demonstrated maturation from an acute effector T cell (TAEFF) to an effector memory T cell (TEM) phenotype with progressive enrichment of KLRG1+CD57+CD272 cells into memory. CMV-specific KLRG1+ TAEFF were capable of in vitro proliferation that diminished upon acquisition of CD57, whereas only KLRG1+ expression correlated with T-bet expression and effector function. In contrast to blood TAEFF, lung mucosal TAEFF demonstrated reduced KLRG1/T-bet expression but similar CD57 levels. Additionally, increased KLRG1+TAEFF were associated with early immune viral control following primary infection. To our knowledge, our findings provide new insights into the roles of KLRG1 and CD57 expression in human T cells, forming the basis for a refined model of CD8+ T cell differentiation during CMV infection. [ABSTRACT FROM AUTHOR]

Immune senescence is a significant contributor to health problems in the developed world and may be accelerated by chronic viral infections. To date, there have been few studies of immune function in healthy older people in sub-Saharan Africa. We assessed T cell and B cell phenotypes and immune responses to CMV, EBV, and influenza virus in Malawians aged 20-69 y. Notably, the proportion of naive (CCR7+CD45RA+) CD4 and CD8 T cells was only 14% of the lymphoid repertoire even in donors aged under 30 y but did not decrease further with age. A small increase in the late differentiated (CD27-CD28-) CD8 T cell subpopulation was observed in older donors but the CD4/CD8 T cell ratio remained stable in all age groups. Interestingly, the regulatory (CD25hiFOXP3hi) T cell subpopulation was small in all age groups, and we observed no age-associated accumulation of cells expressing the senescence- and exhaustion-associated markers CD57 and PD-1. We assessed functional T cell responses to mitogenic and viral antigenic stimulation by the expression of CD154, IFN-γ, TNF-α, IL-2, and IL-17 and proliferation. All responses were robust across the life course, although we observed an age-associated shift from IFN-γ to TNF-α in the response to EBV. In summary, we found the naive T cell subpopulation of young adult Malawians was smaller than in their contemporaries in high-income settings but remains stable thereafter and that lymphocyte function is retained across the life course. These observations indicate that studies of the genetic and environmental factors influencing immune function in different environments may provide insights into minimizing immune ageing. [ABSTRACT FROM AUTHOR]

Virus infection triggers large-scale changes in the phenotype and function of naive CD8+ T cells, resulting in the generation of effector and memory T cells that are then critical for immune clearance. The T-BOX family of transcription factors (TFs) are known to play a key role in T cell differentiation, with mice deficient for the TF T-BET (encoded by Tbx21) unable to generate optimal virus-specific effector responses. Although the importance of T-BET in directing optimal virus-specific T cell responses is accepted, the precise timing and molecular mechanism of action remains unclear. Using a mouse model of influenza A virus infection, we demonstrate that although T-BET is not required for early CD8+ T cell activation and cellular division, it is essential for early acquisition of virus-specific CD8+ T cell function and sustained differentiation and expansion. Whole transcriptome analysis at this early time point showed that Tbx21 deficiency resulted in global dysregulation in early programming events with inappropriate lineage-specific signatures apparent with alterations in the potential TF binding landscape. Assessment of histone posttranslational modifications within the Ifng locus demonstrated that Tbx21 -/- CD8+ T cells were unable to activate "poised" enhancer elements compared with wild-type CD8+ T cells, correlating with diminished Ifng transcription. In all, these data support a model whereby T-BET serves to promote appropriate chromatin remodeling at specific gene loci that underpins appropriate CD8+ T cell lineage-specific commitment and differentiation. [ABSTRACT FROM AUTHOR]

Adipose regulatory T cells (aTregs) have emerged as critical cells for the control of local and systemic inflammation. In this study, we show a distinctive role for the transcriptional regulator Id2 in the differentiation, survival, and function of aTregs in mice. Id2 was highly expressed in aTregs compared with high Id3 expression in lymphoid regulatory T cells (Tregs). Treg-specific deletion of Id2 resulted in a substantial decrease in aTregs, whereas Tregs in the spleen and lymph nodes were unaffected. Additionally, loss of Id2 resulted in decreased expression of aTreg-associated markers, including ST2, CCR2, KLRG1, andGATA3. Gene expression analysis revealed that Id2 expression was essential for the survival of aTregs, and loss of Id2 increased cell death in aTregs due to increased Fas expression. Id2-mediated aTreg depletion resulted in increased systemic inflammation, increased inflammatory macrophages and CD8+ effector T cells, and loss of glucose tolerance under standard diet conditions. Thus, we reveal an unexpected and novel function for Id2 in mediating differentiation, survival, and function of aTregs that when lost result in increased metabolic perturbation. [ABSTRACT FROM AUTHOR]

The transcription factor Helios is expressed in a large percentage of Foxp3+ regulatory T (Treg) cells and is required for the maintenance of their suppressive phenotype, as mice with a selective deficiency of Helios in Treg cells spontaneously develop autoimmunity. However, mice with a deficiency of Helios in all T cells do not exhibit autoimmunity, despite the defect in the suppressor function of their Treg cell population, suggesting that Helios also functions in non-Treg cells. Although Helios is expressed in a small subset of CD4+Foxp3- and CD8+ T cells and its expression is upregulated upon T cell activation, its function in non-Treg cells remains unknown. To examine the function of Helios in CD4+Foxp3- T cells, we transferred Helios-sufficient or -deficient naive CD4+Foxp3- TCR transgenic T cells to normal recipients and examined their capacity to respond to their cognate Ag. Surprisingly, Helios-deficient CD4+ T cells expanded and differentiated into Th1 or Th2 cytokine-producing effectors in a manner similar to wild-type TCR transgenic CD4+ T cells. However, the primed Helios-deficient cells failed to expand upon secondary challenge with Ag. The tolerant state of the Helios-deficient memory T cells was not cell-intrinsic but was due to a small population of Helios-deficient naive T cells that had differentiated into Ag-specific peripheral Treg cells that suppressed the recall response in an Ag-specific manner. These findings demonstrate that Helios plays a role in the determination of CD4+ T cell fate. [ABSTRACT FROM AUTHOR]

T follicular helper (Tfh) cells are essential for germinal center B cell responses. The molecular mechanism underlying the initial Tfh cell differentiation, however, is still incompletely understood. In this study, we show that in vivo, despite enhanced non-Tfh cell effector functions, the deletion of transcription factor Bach2 results in preferential Tfh cell differentiation. Mechanistically, the deletion of Bach2 leads to the induction of CXCR5 expression even before the upregulation of Ascl2. Subsequently, we have identified a novel regulatory element in the murine CXCR5 locus that negatively regulates CXCR5 promoter activities in a Bach2-dependent manner. Bach2 deficiency eventually results in a collapsed CD4+ T cell response with severely impaired CD4+ T cell memory, including Tfh cell memory. Our results demonstrate that Bach2 critically regulates Tfh cell differentiation and CD4+ T cell memory. [ABSTRACT FROM AUTHOR]

CD4+ T cells have been observed to acquire APC-derived membrane and membrane-associated molecules through trogocytosis in diverse immune settings. Despite this, the consequences of trogocytosis on the recipient T cell remain largely unknown. We previously reported that trogocytosed molecules on CD4+ T cells engage their respective surface receptors, leading to sustained TCR signaling and survival after APC removal. Using peptide-pulsed bone marrow-derived dendritic cells and transfected murine fibroblasts expressing antigenic MHC:peptide complexes as APC, we show that trogocytosis-positive CD4+ T cells display effector cytokines and transcription factor expression consistent with a TH2 phenotype. In vitro-polarized TH2 cells were found to be more efficient at performing trogocytosis than TH1 or nonpolarized CD4+ cells, whereas subsequent trogocytosis-mediated signaling induced TH2 differentiation in polarized TH1 and nonpolarized cells. Trogocytosis-positive CD4+ T cells generated in vivo also display a TH2 phenotype in both TCR-transgenic and wild-type models. These findings suggest that trogocytosismediated signaling impacts CD4+ T cell differentiation and effector cytokine production and may play a role in augmenting or shaping a TH2-dominant immune response. [ABSTRACT FROM AUTHOR]

Lymphoid specification is the process by which hematopoietic stem cells (HSCs) and their progeny become restricted to differentiation through the lymphoid lineages. The basic helix-loop-helix transcription factors E2A and Lyl1 form a complex that promotes lymphoid specification. In this study, we demonstrate that Tal1, a Lyl1-related basic helix-loop-helix transcription factor that promotes T acute lymphoblastic leukemia and is required for HSC specification, erythropoiesis, and megakaryopoiesis, is a negative regulator of murine lymphoid specification. We demonstrate that Tal1 limits the expression of multiple E2A target genes in HSCs and controls the balance of myeloid versus T lymphocyte differentiation potential in lymphomyeloid-primed progenitors. Our data provide insight into the mechanisms controlling lymphocyte specification and may reveal a basis for the unique functions of Tal1 and Lyl1 in T acute lymphoblastic leukemia. [ABSTRACT FROM AUTHOR]

Naive CD4+ T lymphocytes differentiate into various Th cell subsets following TCR binding to microbial peptide:MHC class II (p:MHCII) complexes on dendritic cells (DCs). The affinity of the TCR interaction with p:MHCII plays a role in Th differentiation by mechanisms that are not completely understood. We found that low-affinity TCRs biased mouse naive T cells to become T follicular helper (Tfh) cells, whereas higher-affinity TCRs promoted the formation of Th1 or Th17 cells. We explored the basis for this phenomenon by focusing on IL-2R signaling, which is known to promote Th1 and suppress Tfh cell differentiation. SIRPα+ DCs produce abundant p:MHCII complexes and consume IL-2, whereas XCR1+ DCs weakly produce p:MHCII but do not consume IL-2. We found no evidence, however, of preferential interactions between Th1 cell-prone, high-affinity T cells and XCR1+ DCs or Tfh cell-prone, low-affinity T cells and SIRPα+ DCs postinfection with bacteria expressing the peptide of interest. Rather, high-affinity T cells sustained IL-2R expression longer and expressed two novel Th cell differentiation regulators, Eef1e1 and Gbp2, to a higher level than low-affinity T cells. These results suggest that TCR affinity does not influence Th cell differentiation by biasing T cell interactions with IL-2-consuming DCs, but instead, directly regulates genes in naive T cells that control the differentiation process. [ABSTRACT FROM AUTHOR]

The article offers information on an essential function for the autoimmune regulator (Aire) gene in establishing tolerance to tissue-restricted self-antigens during T cell differentiation in the thymus rightfully belongs to this foundational body of work. It discusses tolerance might be learned in the thymus and that immunological tolerance was equivalent to an "immunological thymectomy" with negative selection of a thymic population of self-reactive immune cells.

The article offers information on several immunology findings. Topics include transcription factor retinoic acid receptor– related orphan receptor a (RORa) required for development of innate lymphoid 2 cells (ILC2); and highly polymorphic MHC proteins, monomorphic molecules such as MR1, CD1d, and CD1b present Ags to T cells with specieswide TCR motifs.

At mucosal sites such as the intestine, the immune system launches robust immunity against invading pathogens while maintaining a state of tolerance to commensal flora and ingested food Ags. The molecular mechanisms underlying this phenomenon remain poorly understood. In this study, we report that signaling by GPR81, a receptor for lactate, in colonic dendritic cells and macrophages plays an important role in suppressing colonic inflammation and restoring colonic homeostasis. Genetic deletion of GPR81 in mice led to increased Th1/Th17 cell differentiation and reduced regulatory T cell differentiation, resulting in enhanced susceptibility to colonic inflammation. This was due to increased production of proinflammatory cytokines (IL-6, IL-1b, and TNF-α) and decreased expression of immune regulatory factors (IL-10, retinoic acid, and IDO) by intestinal APCs lacking GPR81. Consistent with these findings, pharmacological activation of GPR81 decreased inflammatory cytokine expression and ameliorated colonic inflammation. Taken together, these findings identify a new and important role for the GPR81 signaling pathway in regulating immune tolerance and colonic inflammation. Thus, manipulation of the GPR81 pathway could provide novel opportunities for enhancing regulatory responses and treating colonic inflammation. [ABSTRACT FROM AUTHOR]

Short-chain fatty acids (SCFAs), the end products of dietary fiber, influence the immune system. Moreover, during pregnancy the maternal microbiome has a great impact on the development of the offspring's immune system. However, the exact mechanisms by which maternal SCFAs during pregnancy and lactation influence the immune system of offspring are not fully understood. We investigated the molecular mechanisms underlying regulatory T cell (Treg) differentiation in offspring regulated by a maternal high fiber diet (HFD). Plasma levels of SCFAs in offspring from HFD-fed mice were higher than in those from no fiber diet--fed mice. Consequently, the offspring from HFD-fed mice had higher frequencies of thymic Treg (tTreg) and peripheral Tregs. We found that the offspring of HFD-fed mice exhibited higher autoimmune regulator (Aire) expression, a transcription factor expressed in the thymic microenvironment, suggesting SCFAs promote tTreg differentiation through increased Aire expression. Notably, the receptor for butyrate, G protein--coupled receptor 41 (GPR41), is highly expressed in the thymic microenvironment and Aire expression is not increased by stimulation with butyrate in GPR41-deficient mice. Our studies highlight the significance of SCFAs produced by a maternal HFD for Treg differentiation in the thymus of offspring. Given that Aire expression is associated with the induction of tTregs, the maternal microbiome influences Treg differentiation in the thymus of offspring through GPR41-mediated Aire expression. The [ABSTRACT FROM AUTHOR]

In response to acute viral infection, activated naive T cells give rise to effector T cells that clear the pathogen and memory T cells that persist long-term and provide heightened protection. T cell factor 1 (Tcf1) is essential for several of these differentiation processes. Tcf1 is expressed in multiple isoforms, with all isoforms sharing the same HDAC and DNA-binding domains and the long isoforms containing a unique N-terminal β-catenin-interacting domain. In this study, we specifically ablated Tcf1 long isoforms in mice, while retaining expression of Tcf1 short isoforms. During CD8+ T cell responses, Tcf1 long isoforms were dispensable for generating cytotoxic CD8+ effector T cells and maintaining memory CD8+ T cell pool size, but they contributed to optimal maturation of central memory CD8+ T cells and their optimal secondary expansion in a recall response. In contrast, Tcf1 long isoforms were required for differentiation of T follicular helper (TFH) cells, but not TH1 effectors, elicited by viral infection. Although Tcf1 short isoforms adequately supported Bcl6 and ICOS expression in TFH cells, Tcf1 long isoforms remained important for suppressing the expression of Blimp1 and TH1-associated genes and for positively regulating Id3 to restrain germinal center TFH cell differentiation. Furthermore, formation of memory TH1 and memory TFH cells strongly depended on Tcf1 long isoforms. These data reveal that Tcf1 long and short isoforms have distinct, yet complementary, functions and may represent an evolutionarily conserved means to ensure proper programming of CD8+ and CD4+ T cell responses to viral infection. [ABSTRACT FROM AUTHOR]

Transcription factor retinoid acid-related orphan receptor (ROR)γt transcriptionally regulates the genes required for differentiation of Th17 cells that mediate both protective and pathogenic immunity. However, little is known about the function of posttranslational modifications in the regulation of RORγt activity. Mass spectrometric analysis of immunoprecipitated RORγt from Th17 cells identified multiple phosphorylation sites. Systematic mutation analysis of the identified phosphorylation sites found that phosphorylation of S376 enhances whereas phosphorylation of S484 inhibits Th17 differentiation. IkB kinase (IKK)α binds and phosphorylates RORγt at S376 but not S484. Knockdown of IKKα, dominant-negative IKKα, and RORγt mutants incapable of interacting with IKKα all decrease Th17 differentiation. Furthermore, nonphosophorylatable RORγt mutant (S376A) impairs whereas phosphomimetic mutant (S376E) stimulates Th17 differentiation independent of IKKα. Therefore, IKKα-dependent phosphorylation of S376 stimulated whereas IKKα-independent phosphorylation of S484 inhibited RORγt function in Th17 differentiation. [ABSTRACT FROM AUTHOR]

Neonatal and adult T cells differ in their effector functions. Although it is known that cell-intrinsic differences in mature T cells contribute to this phenomenon, the factors involved remain unclear. Given emerging evidence that the binding strength of a TCR for self-peptide presented by MHC (self-pMHC) impacts T cell function, we sought to determine whether altered thymic selection influences the self-reactivity of the TCR repertoire during ontogeny. We found that conventional and regulatory T cell subsets in the thymus of neonates and young mice expressed higher levels of cell surface CD5, a surrogate marker for TCR avidity for self-pMHC, as compared with their adult counterparts, and this difference in self-reactivity was independent of the germline bias of the neonatal TCR repertoire. The increased binding strength of the TCR repertoire for self-pMHC in neonates was not solely due to reported defects in clonal deletion. Rather, our data suggest that thymic selection is altered in young mice such that thymocytes bearing TCRs with low affinity for self-peptide are not efficiently selected into the neonatal repertoire, and stronger TCR signals accompany both conventional and regulatory T cell selection. Importantly, the distinct levels of T cell self-reactivity reflect physiologically relevant differences based on the preferential expansion of T cells from young mice to fill a lymphopenic environment. Therefore, differences in thymic selection in young versus adult mice skew the TCR repertoire, and the relatively higher selfreactivity of the T cell pool may contribute to the distinct immune responses observed in neonates. [ABSTRACT FROM AUTHOR]

Innate regulation through TLR signaling has been shown to be important for promoting T cell subset development and function. However, limited information is known about whether differential TLR signaling can selectively inhibit Th17 and/or Th1 cells, which are important for controlling excessive inflammation and autoimmune responses. In this article, we demonstrate that activation of TLR7 signaling in T cells can inhibit Th17 cell differentiation from naive T cells and IL-17 production in established Th17 cells. We further report that downregulation of STAT3 signaling is responsible for TLR7-mediated inhibition of Th17 cells due to induction of suppressor of cytokine signaling 3 and 5. TLR7-mediated suppression of Th17 cells does not require dendritic cell involvement. In addition, we show that TLR7 signaling can suppress Th1 cell development and function through a mechanism different from Th17 cell suppression. Importantly, our complementary in vivo studies demonstrate that treatment with the TLR7 ligand imiquimod can inhibit Th1 and Th17 cells, resulting in the prevention of, and an immunotherapeutic reduction in, experimental autoimmune encephalomyelitis. These studies identify a new strategy to manipulate Th17/Th1 cells through TLR7 signaling, with important implications for successful immunotherapy against autoimmune and inflammatory diseases. [ABSTRACT FROM AUTHOR]

Th17 cells play a critical role in autoimmune diseases, including multiple sclerosis and its animal model, experimental autoimmune encephalomyelitis. Response gene to complement (RGC)-32 is a cell cycle regulator and a downstream target of TGF-β that mediates its profibrotic activity. In this study, we report that RGC-32 is preferentially upregulated during Th17 cell differentiation. RGC-32-/- mice have normal Th1, Th2, and regulatory T cell differentiation but show defective Th17 differentiation in vitro. The impaired Th17 differentiation is associated with defects in IFN regulatory factor 4, B cell-activating transcription factor, retinoic acid-related orphan receptor γt, and SMAD2 activation. In vivo, RGC-32-/- mice display an attenuated experimental autoimmune encephalomyelitis phenotype accompanied by decreased CNS inflammation and reduced frequency of IL-17- and GM-CSF-producing CD4+ T cells. Collectively, our results identify RGC-32 as a novel regulator of Th17 cell differentiation in vitro and in vivo and suggest that RGC-32 is a potential therapeutic target in multiple sclerosis and other Th17-mediated autoimmune diseases. [ABSTRACT FROM AUTHOR]

Tissue-resident memory T cells (TRM) have been shown to afford superior protection against infection, particularly against pathogens that enter via the epithelial surfaces of the body. Although TRM are often concentrated at sites of prior infection, it has been shown that TRM can disseminate throughout the body. We examined the relative effectiveness of global versus targeted CD8+ TRM lodgment in skin. The site of initial T cell priming made little difference to skin lodgement, whereas local inflammation and Ag recognition enhanced TRM accumulation and retention. Disseminated TRM lodgment was seen with the skin, but required multiple exposures to Ag and was inferior to targeted strategies. As a consequence, active recruitment by inflammation or infection resulted in superior TRM numbers and maximal protection against infection. Overall, these results highlight the potency of localized TRM deposition as a means of pathogen control as well as demonstrating the limitations of global TRM lodgment. [ABSTRACT FROM AUTHOR]

Although CD31 expression on human thymocytes has been reported, a detailed analysis of CD31 expression at various stages of T cell development in the human thymus is missing. In this study, we provide a global picture of the evolution of CD31 expression from the CD34+ hematopoietic precursor to the CD45RA+ mature CD4+ and CD8+ single-positive (SP) T cells. Using nine-color flow cytometry, we show that CD31 is highly expressed on CD34+ progenitors and stays high until the early double-positive stage (CD3-CD4+CD8α+β-). After β-selection, CD31 expression levels become low to undetectable. CD31 expression then increases and peaks on CD3highCD4+CD8+ double-positive thymocytes. However, following positive selection, CD31 expression differs dramatically between CD4+ and CD8+ lineages: homogeneously high on CD8 SP but lower or negative on CD4 SP cells, including a subset of CD45RA+CD31- mature CD4+ thymocytes. CD31 expression on TCRγδ thymocytes is very similar to that of CD4 SP cells. Remarkably, there is a substantial subset of semimature (CD45RA-) CD4 SP thymocytes that lack CD31 expression. Moreover, FOXP3+ and ICOS+ cells are overrepresented in this CD31- subpopulation. Despite this CD31-CD45RA- subpopulation, most egress-capable mature CD45RA+ CD4 SP thymocytes express CD31. The variations in CD31 expression appear to coincide with three major selection processes occurring during thymopoiesis: β-selection, positive selection, and negative selection. Considering the ability of CD31 to modulate the TCR's activation threshold via the recruitment of tyrosine phosphatases, our results suggest a significant role for CD31 during T cell development. [ABSTRACT FROM AUTHOR]

Nucleotide oligomerization domain-like receptor X1 (NLRX1) has been implicated in viral response, cancer progression, and inflammatory disorders; however, its role as a dual modulator of CD4+ T cell function and metabolism has not been defined. The loss of NLRX1 results in increased disease severity, populations of Th1 and Th17 cells, and inflammatory markers (IFN-γ, TNF-α, and IL-17) in mice with dextran sodium sulfate-induced colitis. To further characterize this phenotype, we used in vitro CD4+ T cell-differentiation assays and show that NLRX1-deficient T cells have a greater ability to differentiate into an inflammatory phenotype and possess greater proliferation rates. Further, NLRX1-/- cells have a decreased responsiveness to immune checkpoint pathways and greater rates of lactate dehydrogenase activity. When metabolic effects of the knockout are impaired, NLRX1-deficient cells do not display significant differences in differentiation or proliferation. To confirm the role of NLRX1 specifically in T cells, we used an adoptive-transfer model of colitis. Rag2-/- mice receiving NLRX1-/- naive or effector T cells experienced increased disease activity and effector T cell populations, whereas no differences were observed between groups receiving wildtype or NLRX1-/- regulatory T cells. Metabolic effects of NLRX1 deficiency are observed in a CD4-specific knockout of NLRX1 within a Citrobacter rodentium model of colitis. The aerobic glycolytic preference in NLRX1-/- effector T cells is combined with a decreased sensitivity to immunosuppressive checkpoint pathways to provide greater proliferative capabilities and an inflammatory phenotype bias leading to increased disease severity. [ABSTRACT FROM AUTHOR]

Regulatory T (Treg) cells establish tolerance, prevent inflammation at mucosal surfaces, and regulate immunopathology during infectious responses. Recent studies have shown that Delta-like ligand 4 (Dll4) was upregulated on APC after respiratory syncytial virus (RSV) infection, and its inhibition leads to exaggerated immunopathology. In the present study, we outline the role of Dll4 in Treg cell differentiation, stability, and function in RSV infection. We found that Dll4 was expressed on CD11b+ pulmonary dendritic cells in the lung and draining lymph nodes in wild-type BALB/c mice after RSV infection. Dll4 neutralization exacerbated RSV-induced disease pathology, mucus production, group 2 innate lymphoid cell infiltration, IL-5 and IL-13 production, as well as IL-17A+ CD4 T cells. Dll4 inhibition decreased the abundance of CD62LhiCD44loFoxp3+ central Treg cells in draining lymph nodes. The RSV-induced disease was accompanied by an increase in Th17-like effector phenotype in Foxp3+ Treg cells and a decrease in granzyme B expression after Dll4 blockade. Finally, Dll4-exposed induced Treg cells maintained the CD62LhiCD44lo central Treg cell phenotype, had increased Foxp3 expression, became more suppressive, and were resistant to Th17 skewing in vitro. These results suggest that Dll4 activation during differentiation sustained Treg cell phenotype and function to control RSV infection. [ABSTRACT FROM AUTHOR]

Epigenetic mechanisms play a critical role during differentiation of T cells by contributing to the formation of stable and heritable transcriptional patterns. To better understand the mechanisms of memory maintenance in CD8+ T cells, we performed genome-wide analysis of DNA methylation, histone marking (acetylated lysine 9 in histone H3 and trimethylated lysine 9 in histone), and gene-expression profiles in naive, effector memory (EM), and terminally differentiated EM (TEMRA) cells. Our results indicate that DNA demethylation and histone acetylation are coordinated to generate the transcriptional program associated with memory cells. Conversely, EM and TEMRA cells share a very similar epigenetic landscape. Nonetheless, the TEMRA transcriptional program predicts an innate immunity phenotype associated with genes never reported in these cells, including several mediators of NK cell activation (VAV3 and LYN) and a large array of NK receptors (e.g., KIR2DL3, KIR2DL4, KIR2DL1, KIR3DL1, KIR2DS5). In addition, we identified up to 161 genes that encode transcriptional regulators, some of unknown function in CD8+ T cells, and that were differentially expressed in the course of differentiation. Overall, these results provide new insights into the regulatory networks involved in memory CD8+ T cell maintenance and T cell terminal differentiation. [ABSTRACT FROM AUTHOR]

Tissue-resident memory T (TRM) cells, a population of noncirculating memory T cells, are one of the essential components of immunological memory in both mouse and human. Although CD69+CD103+ TRM cells represent a major TRM cell population in barrier tissues including the mucosal surface and the skin, CD69+CD103- TRM cells dominate most nonbarrier tissues, such as the kidney. TGF-β is required for the differentiation of CD69+CD103+ TRM cells in barrier tissues. However, the developmental control of CD69+CD103- TRM cells in nonbarrier tissues remains largely unknown and the involvement of TGF-β signaling is less clear. In this study we demonstrated that TGF-β promoted the formation of kidney-resident T cells via enhancing the tissue entry of effector T cells. Mechanistically, TGF-β enhanced E- and P-selectin and inflammatory chemokine-mediated extravasation of effector T cells. Thus TGF-β controls the first developmental checkpoint of TRM cell differentiation in nonbarrier tissues. [ABSTRACT FROM AUTHOR]

Concurrent helminth infection potently inhibits T cell immunity; however, whether helminthes prevent T cell priming or skew clonal recruitment and effector differentiation is not known. Using coinfection with two natural mouse pathogens, Heligsomosoides polygyrus and Toxoplasma gondii, to investigate the negative impact of helminthes on the CD8 T cell response, we demonstrate helminth-induced suppression of IL-12-dependent differentiation of killer-like receptor G1+ effector CD8 T cells and IFN-γ production. Nevertheless, reversal of helminth suppression of the innate IL-12 response of CD8α+ dendritic cells, which occurred in STAT6-deficient mice, was not sufficient to normalize CD8 T cell differentiation. Instead, a combined deficiency in IL-4 and IL-10 was required to reverse the negative effects of helminth coinfection on the CD8 T cell response. Monoclonal T. gondii-specific CD8 T cells adoptively transferred into coinfected mice recapitulated the spectrum of helminth-induced effects on the polyclonal CD8 T response, indicating the lack of requirement for clonal skewing. [ABSTRACT FROM AUTHOR]

Adenosine A2a receptor (A2aR) signaling acts as a barrier to autoimmunity by promoting anergy, inducing regulatory T cells, and inhibiting effector T cells. However, in vivo effects of A2aR signaling on polyclonal CD4 T cells during a primary response to foreign Ag has yet to be determined. To address this problem, we immunized mice with peptide Ag 2W1S coupled to PE in CFA and treated with the selective A2aR agonist CGS-21680 (CGS). 2W1S:I-Ab-specific tetramer-binding CD4 T cells did not become anergic or differentiate into Foxp3+ regulatory T cells. Additionally, CGS treatment did not inhibit Th1 or Th17 differentiation. However, CGS did abrogate germinal center T follicular helper cells, and blunted PE-specific germinal center B cell responses. The use of A2aR-deficient CD4 T cells established that this CGS effect was T cell intrinsic. Therefore, this study has identified a unique role for A2aRs in regulating CD4 T cell differentiation during vaccination. [ABSTRACT FROM AUTHOR]

Expression of programmed death 1 (PD-1) on CD8 T cells promotes T cell exhaustion during chronic Ag exposure. During acute infections, PD-1 is transiently expressed and has the potential to modulate CD8 T cell memory formation. Conserved region C (CR-C), a promoter proximal cis-regulatory element that is critical to PD-1 expression in vitro, responds to NFATc1, FoxO1, and/or NF-kB signaling pathways. Here, a CR-C knockout mouse was established to determine its role on PD-1 expression and the corresponding effects on T cell function in vivo. Deletion of CR-C decreased PD-1 expression on CD4 T cells and Ag-specific CD8 T cells during acute and chronic lymphocytic choriomeningitis virus challenges, but did not affect the ability to clear an infection. Following acute lymphocytic choriomeningitis virus infection, memory CD8 T cells in the CR-C knockout mouse were formed in greater numbers, were more functional, and were more effective at responding to a melanoma tumor than wild-type memory cells. These data implicate a critical role for CR-C in governing PD-1 expression, and a subsequent role in guiding CD8 T cell differentiation. The data suggest the possibility that titrating PD-1 expression during CD8 T cell activation could have important ramifications in vaccine development and clinical care. [ABSTRACT FROM AUTHOR]

Immune tolerance between the fetus and mother represents an active process by which the developing fetus must not mount immune responses to noninherited Ags on chimeric maternal cells that reside in fetal tissue. This is, in part, mediated by the suppressive influence of CD4+FOXP3+CD25+ regulatory T cells (Tregs). Fetal secondary lymphoid organs have an increased frequency of Tregs and, as compared with adult T cells, fetal naive CD4+ T cells exhibit a strong predisposition to differentiate into Tregs when stimulated. This effect is mediated by the TCR and TGF-β pathways, and fetal T cells show significantly increased Treg differentiation in response to anti-CD3 and TGF-β stimulation. Naive fetal T cells also exhibit increased signaling through the TGF-β pathway, with these cells demonstrating increased expression of the signaling mediators TGF-βRI, TGF-βRIII, and SMAD2, and higher levels of SMAD2/SMAD3 phosphorylation. Increased fetal Treg differentiation is mediated by the RNA-binding protein Lin28b, which is overexpressed in fetal T cells as compared with adult cells. When Lin28b expression is decreased in naive fetal T cells, they exhibit decreased Treg differentiation that is associated with decreased TGF-β signaling and lowered expression of TGF-βRI, TGF-βRIII, and SMAD2. Lin28b regulates the maturation of let-7 microRNAs, and these TGF-β signaling mediators are let-7 targets. We hypothesize that loss of Lin28b expression in fetal T cells leads to increased mature let-7, which causes decreased expression of TGF-βRI, TGF-βRIII, and SMAD2 proteins. A reduction in TGF-β signaling leads to reduced Treg numbers. [ABSTRACT FROM AUTHOR]

The use of nondepleting Abs specific for CD4 and CD8 is an effective strategy to tolerize CD4+ and CD8+ T cells in a tissue-specific manner. We reported that coreceptor therapy reverses diabetes in new onset NOD mice. A striking feature of coreceptor-induced remission is the purging of T cells from the pancreatic lymph nodes (PLN) and islets of NOD mice. Evidence indicates that Abs binding to the coreceptors promotes T cell egress from these tissues. The present study examined how coreceptor therapy affects the migration of CD4+ T cells residing in the PLN of NOD mice. Anti-CD4 Ab treatment resulted in an increased frequency of PLN but not splenic CD4+ T cells that exhibited a polarized morphology consistent with a migratory phenotype. Furthermore, PLN CD4+ T cells isolated from anti-CD4 versus control Ab-treated animals displayed increased in vitro chemotaxis to chemoattractants such as sphingosine-1-phosphate and CXCL12. Notably, the latter was dependent on activation of the small Rho GTPases Rac1 and Rac2. Rac1 and Rac2 activation was increased in Ab-bound CD4+ T cells from the PLN but not the spleen, and knockdown of Rac expression blocked the heightened reactivity of Ab-bound PLN CD4+ T cells to CXCL12. Interestingly, Rac1 and Rac2 activation was independent of Rac guanine nucleotide exchange factors known to regulate T cell activity. Therefore, Ab binding to CD4 initiates a novel pathway that involves inflammation-dependent activation of Rac and establishment of altered T cell migratory properties. [ABSTRACT FROM AUTHOR]

We tested, in rhesus macaques, the effects of a 500-fold range of an admixed recombinant modified vaccinia Ankara (MVA) expressing rhesus GM-CSF (MVA/GM-CSF) on the immunogenicity and protection elicited by an MVA/SIV macaque 239 vaccine. High doses of MVA/GM-CSF did not affect the levels of systemic envelope (Env)-specific Ab, but it did decrease the expression of the gut-homing receptor α4β7 on plasmacytoid dendritic cells (p < 0.01) and the magnitudes of Env-specific IgA (p = 0.01) and IgG (p < 0.05) in rectal secretions. The protective effect of the vaccine was evaluated using 12 weekly rectal challenges in rhesus macaques subgrouped by tripartite motif-containing protein 5α (TRIM5α) genotypes that are restrictive or permissive for infection by the challenge virus SIVsmE660. Eight of nine TRIM5α-restrictive animals receiving no or the lowest dose (1 × 105 PFU) of MVA/GM-CSF resisted all 12 challenges. In the comparable TRIM5α-permissive group, only 1 of 12 animals resisted all 12 challenges. In the TRIM5α-restrictive animals, but not in the TRIM5α-permissive animals, the number of challenges to infection directly correlated with the magnitudes of Env-specific rectal IgG (r = +0.6) and IgA (r = +0.6), the avidity of Env-specific serum IgG (r = +0.5), and Ab dependent cell-mediated virus inhibition (r = +0.6). Titers of neutralizing Ab did not correlate with protection. We conclude that 1) protection elicited by MVA/SIVmac239 is strongly dependent on the presence of TRIM5α restriction, 2) nonneutralizing Ab responses contribute to protection against SIVsmE660 in TRIM5α-restrictive animals, and 3) high doses of codelivered MVA/GM-CSF inhibit mucosal Ab responses and the protection elicited by MVA expressing noninfectious SIV macaque 239 virus-like particles. [ABSTRACT FROM AUTHOR]

Periodontitis is a chronic oral inflammatory disease affecting one in five individuals that can lead to tooth loss. CD4+ Th cells activated by a microbial biofilm are thought to contribute to the destruction of alveolar bone surrounding teeth by influencing osteoclastogenesis through IL-17A and receptor activator for NF-κB ligand effects. The relative roles of mucosal Ag presentation cells in directing Th cell immune responses against oral pathogens and their contribution to destruction of alveolar bone remain unknown. We tested the contribution of mucosal Langerhans cells (LCs) to alveolar bone homeostasis in mice following oral colonization with a well-characterized human periodontal pathogen, Porphyromonas gingivalis. We found that oral mucosal LCs did not protect from or exacerbate crestal alveolar bone destruction but were responsible for promoting differentiation of Th17 cells specific to P. gingivalis. In mice lacking LCs the Th17 response was suppressed and a Th1 response predominated. Bypassing LCs with systemic immunization of P. gingivalis resulted in a predominantly P. gingivalis-specific Th1 response regardless of whether LCs were present. Interestingly, we find that in vivo clonal expansion of P. gingivalis-specific Th cells and induced regulatory T cells does not depend on mucosal LCs. Furthermore, destruction of crestal alveolar bone induced by P. gingivalis colonization occurred regardless of the presence of mucosal LCs or P. gingivalis-specific Th17 cells. Our data indicate that both LCs and Th17 cells are redundant in contributing to alveolar bone destruction in a murine model of periodontitis. [ABSTRACT FROM AUTHOR]

Recent data illustrate a key role for the transcriptional regulator bric-a-brac, tramtrack, and broad complex and cap'n'collar homology (Bach)2 in orchestrating T cell differentiation and function. Although Bach2 has a well-described role in B cell differentiation, emerging data show that Bach2 is a prototypical member of a novel class of transcription factors that regulates transcriptional activity in T cells at super-enhancers, or regions of high transcriptional activity. Accumulating data demonstrate specific roles for Bach2 in favoring regulatory T cell generation, restraining effector T cell differentiation, and potentiating memory T cell development. Evidence suggests that Bach2 regulates various facets of T cell function by repressing other key transcriptional regulators such as B lymphocyte-induced maturation protein 1. In this review, we examine our present understanding of the role of Bach2 in T cell function and highlight the growing evidence that this transcriptional repressor functions as a key regulator involved in maintenance of T cell quiescence, T cell subset differentiation, and memory T cell generation. [ABSTRACT FROM AUTHOR]

The scaffold molecule POSH is crucial for the regulation of proliferation and effector function in CD8+ T cells. However, its role in CD4 1 cells is not known. In this study, we found that disruption of the POSH scaffold complex established a transcriptional profile that strongly skewed differentiation toward Th2, led to decreased survival, and had no effect on cell cycle entry. This is in stark contrast to CD8+ T cells in which POSH regulates cell cycle and does not affect survival. Disruption of POSH in CD4+ T cells resulted in the loss of Takl-dependent activation of JNK1/2 and Takl-mediated survival. However, in CD8+ T cells, POSH regulates only JNK1. Remarkably, each type of T cell had a unique composition of the POSH scaffold complex and distinct posttranslational modifications of POSH. These data indicate that the mechanism that regulates POSH function in CD4+ T cells is different from CD8+ T cells. All together, these data strongly suggest that POSH is essential for the integration of cell-type-specific signals that regulate the differentiation, survival, and function of T cells. [ABSTRACT FROM AUTHOR]

γδ T lymphocytes are programmed into distinct IFN-γ-producing CD27+ (γδ27+) and IL-17-producing CD27- (γδ27-) subsets that play key roles in protective or pathogenic immune responses. Although the signature cytokines are shared with their αβ Th1 (for γδ27+) and Th17 (for γδ27-) cell counterparts, we dissect in this study similarities and differences in the transcriptional requirements of murine effector γδ27+, γδ27-CCR6-, and γδ27-CCR6+ γδ T cell subsets and αβ T cells. We found they share dependence on the master transcription factors T-bet and RORγt for IFN-γ and IL-17 production, respectively. However, Eomes is fully dispensable for IFN-γ production by γδ T cells. Furthermore, the Th17 cell auxiliary transcription factors RORα and BATF are not required for IL-17 production by γδ27- cell subsets. We also show that γδ27- (but not γδ27+) cells become polyfunctional upon IL-1β plus IL-23 stimulation, cosecreting IL-17A, IL-17F, IL-22, GM-CSF, and IFN-γ. Collectively, our in vitro and in vivo data firmly establish the molecular segregation between γδ27+ and γδ27- T cell subsets and provide novel insight on the nonoverlapping transcriptional networks that control the differentiation of effector γδ versus αβ T cell subsets. [ABSTRACT FROM AUTHOR]