The Th1 cell regulatory circuitry is largely conserved between human and mouse.

Gene expression programs controlled by lineage-determining transcription factors are often conserved between species. However, infectious diseases have exerted profound evolutionary pressure, and therefore the genes regulated by immune-specific transcription factors might be expected to exhibit greater divergence. T-bet (Tbx21) is the immune-specific, lineage-specifying transcription factor for T helper type I (Th1) immunity, which is fundamental for the immune response to intracellular pathogens but also underlies inflammatory diseases. We compared T-bet genomic targets between mouse and human CD4+ T cells and correlated T-bet binding patterns with species-specific gene expression. Remarkably, we found that the majority of T-bet target genes are conserved between mouse and human, either via preservation of binding sites or via alternative binding sites associated with transposon-linked insertion. Species-specific T-bet binding was associated with differences in transcription factor-binding motifs and species-specific expression of associated genes. These results provide a genome-wide cross-species comparison of Th1 gene regulation that will enable more accurate translation of genetic targets and therapeutics from pre-clinical models of inflammatory and infectious diseases and cancer into human clinical trials.

The energy sensor AMPK orchestrates metabolic and translational adaptation in expanding T helper cells.

The importance of cellular metabolic adaptation in inducing robust T cell responses is well established. However, the mechanism by which T cells link information regarding nutrient supply to clonal expansion and effector function is still enigmatic. Herein, we report that the metabolic sensor adenosine monophosphate-activated protein kinase (AMPK) is a critical link between cellular energy demand and translational activity and, thus, orchestrates optimal expansion of T cells in vivo. AMPK deficiency did not affect T cell fate decision, activation, or T effector cell generation; however, the magnitude of T cell responses in murine in vivo models of T cell activation was markedly reduced. This impairment was global, as all T helper cell subsets were similarly sensitive to loss of AMPK which resulted in reduced T cell accumulation in peripheral organs and reduced disease severity in pathophysiologically as diverse models as T cell transfer colitis and allergic airway inflammation. T cell receptor repertoire analysis confirmed similar clonotype frequencies in different lymphoid organs, thereby supporting the concept of a quantitative impairment in clonal expansion rather than a skewed qualitative immune response. In line with these findings, in-depth metabolic analysis revealed a decrease in T cell oxidative metabolism, and gene set enrichment analysis indicated a major reduction in ribosomal biogenesis and mRNA translation in AMPK-deficient T cells. We, thus, provide evidence that through its interference with these delicate processes, AMPK orchestrates the quantitative, but not the qualitative, manifestation of primary T cell responses in vivo.

Th1-Polarized, Dengue Virus-Activated Human Mast Cells Induce Endothelial Transcriptional Activation and Permeability.

Dengue virus (DENV), an arbovirus, strongly activates mast cells (MCs), which are key immune cells for pathogen immune surveillance. In animal models, MCs promote clearance of local peripheral DENV infections but, conversely, also promote pathological vascular leakage when widely activated during systemic DENV infection. Since DENV is a human pathogen, we sought to ascertain whether a similar phenomenon could occur in humans by characterizing the products released by human MCs (huMCs) upon direct (antibody-independent) DENV exposure, using the phenotypically mature huMC line, ROSA. DENV did not productively infect huMCs but prompted huMC release of proteases and eicosanoids and induced a Th1-polarized transcriptional profile. In co-culture and trans-well systems, huMC products activated human microvascular endothelial cells, involving transcription of vasoactive mediators and increased monolayer permeability. This permeability was blocked by MC-stabilizing drugs, or limited by drugs targeting certain MC products. Thus, MC stabilizers are a viable strategy to limit MC-promoted vascular leakage during DENV infection in humans.

Pterostilbene reduces colonic inflammation by suppressing dendritic cell activation and promoting regulatory T cell development.

Dendritic cells (DCs) and T cells play important roles in immune regulation, and modulating their function is an approach for developing preventive or therapeutic strategies against immune disorders. Herein, the effect of pterostilbene (PSB) (3',5'-dimethoxy-resveratrol)-a resveratrol-related polyphenol found in blueberries-on immune regulation was evaluated. Using an in vitro co-culture system, PSB was found to exert the strongest inhibitory effect among all tested resveratrol derivatives on DC-mediated T cell proliferation; moreover, PSB treatment decreased the Th1 and Th17 populations and increased the regulatory T cell (Treg) population. Upon co-stimulation with anti-CD3 and anti-CD28 antibodies, PSB inhibited CD4+ T cell proliferation and differentiation into Th1 cells. Additionally, PSB acted on DCs to suppress the lipopolysaccharide-induced transactivation of genes encoding antigen presentation-related molecules and inflammatory cytokines by attenuating the DNA-binding ability of the transcription factor PU.1. Furthermore, PSB promoted DC-mediated Foxp3+ Treg differentiation, and PU.1 knockdown increased DC-induced Treg activity. Oral administration of PSB alleviated the symptoms of dextran sulfate sodium-induced colitis and decreased tumor necrosis factor-α expression in mice. Thus, PSB treatment ameliorates colonic inflammation.

Acetylcholine regulates the development of experimental autoimmune encephalomyelitis via the CD4+ cells proliferation and differentiation.

Purpose of the study: Multiple sclerosis is a CD4+ T cell mediated autoimmune disease characterized by inflammatory demyelination in the central nervous system. Acetylcholine (ACh) has been reported to be released by T lymphocytes and plays as an inflammation and immune regulator through the participation of T cells. However, both attenuated and aggravated effects of ACh in inflammation were found. The aim of this study is to further investigate the role of ACh in experimental autoimmune encephalomyelitis (EAE).Materials and methods: The left cervical vagotomy was performed to inhibit ACh release with the sham-operation as control. ACh in cerebral cortex and splenocytes culture supernatants of EAE mice were determined. Interleukin-6, interferon-γ, interleukin-4 and interleukin-17A in brain and splenocytes culture supernatants were evaluated by enzyme-linked immunosorbent assay. The proportion of CD4+ T cells and subsets were assessed by flow cytometry.Results: Compared with the sham-operation group, improved clinical and pathological parameters as well as decreased interleukin-6, interferon-γ, interleukin-4 and interleukin-17A were found in EAE mice with vagotomy suppressing the ACh. Marked reductions of CD4+ and CD4+ChAT+ cells, as well as significant decrease in Th1 with a bias to Th2 in Th1/Th2 balance and increased ChAT+Th2 proportion in the spleen were also observed in vagotomized mice.Conclusions: These findings emphasize that inhibiting ACh release by vagotomy can ameliorate the exacerbation of EAE through suppressing CD4+ T cells proliferation and regulating the differentiation of Th1, Th2 and Th17.

NF-κB/mTOR/MYC Axis Drives PRMT5 Protein Induction After T Cell Activation via Transcriptional and Non-transcriptional Mechanisms.

Multiple sclerosis is an autoimmune disease of the central nervous system (CNS) mediated by CD4+ T cells and modeled via experimental autoimmune encephalomyelitis (EAE). Inhibition of PRMT5, the major Type II arginine methyltransferase, suppresses pathogenic T cell responses and EAE. PRMT5 is transiently induced in proliferating memory inflammatory Th1 cells and during EAE. However, the mechanisms driving PRMT5 protein induction and repression as T cells expand and return to resting is currently unknown. Here, we used naive mouse and memory mouse and human Th1/Th2 cells as models to identify mechanisms controlling PRMT5 protein expression in initial and recall T cell activation. Initial activation of naive mouse T cells resulted in NF-κB-dependent transient Prmt5 transcription and NF-κB, mTOR and MYC-dependent PRMT5 protein induction. In murine memory Th cells, transcription and miRNA loss supported PRMT5 induction to a lesser extent than in naive T cells. In contrast, NF-κB/MYC/mTOR-dependent non-transcriptional PRMT5 induction played a major role. These results highlight the importance of the NF-κB/mTOR/MYC axis in PRMT5-driven pathogenic T cell expansion and may guide targeted therapeutic strategies for MS.

Cognate Interaction With CD4+ T Cells Instructs Tumor-Associated Macrophages to Acquire M1-Like Phenotype.

The immunosuppressive tumor microenvironment (TME) established by tumor cells, stromal cells and inhibitory immune cells counteracts the function of tumor reactive T cells. Tumor associated macrophages (TAMs) showing functional plasticity contribute to this process as so called M2-like macrophages can suppress the function of effector T cells and promote their differentiation into regulatory T cells (Tregs). Furthermore, tumor antigen specific CD4+ T effector cells can essentially sustain anti-tumoral immune responses as shown for various tumor entities, thus suggesting that cognate interaction between tumor antigen-specific CD4+ Th1 cells and TAMs might shift the intra-tumoral M1/M2 ratio toward M1. This study demonstrates repolarization of M2-like PECs upon MHC II-restricted interaction with tumor specific CD4+ Th1 cells in vitro as shown by extensive gene and protein expression analyses. Moreover, adoptive transfer of OVA-specific OT-II cells into C57BL/6 mice bearing OVA expressing IAb-/- tumors resulted in increased accumulation of M1-like TAMs with enhanced M1 associated gene and protein expression profiles. Thus, this paper highlights a so far underestimated function of the CD4+ Th1/TAM axis in re-conditioning the immunosuppressive tumor microenvironment.

A chemical biology toolbox to study protein methyltransferases and epigenetic signaling.

Protein methyltransferases (PMTs) comprise a major class of epigenetic regulatory enzymes with therapeutic relevance. Here we present a collection of chemical probes and associated reagents and data to elucidate the function of human and murine PMTs in cellular studies. Our collection provides inhibitors and antagonists that together modulate most of the key regulatory methylation marks on histones H3 and H4, providing an important resource for modulating cellular epigenomes. We describe a comprehensive and comparative characterization of the probe collection with respect to their potency, selectivity, and mode of inhibition. We demonstrate the utility of this collection in CD4+ T cell differentiation assays revealing the potential of individual probes to alter multiple T cell subpopulations which may have implications for T cell-mediated processes such as inflammation and immuno-oncology. In particular, we demonstrate a role for DOT1L in limiting Th1 cell differentiation and maintaining lineage integrity. This chemical probe collection and associated data form a resource for the study of methylation-mediated signaling in epigenetics, inflammation and beyond.

A predominant Th1 polarization is present in synovial fluid of end-stage osteoarthritic knee joints: analysis of peripheral blood, synovial fluid and synovial membrane.

Thorough understanding of the complex pathophysiology of osteoarthritis (OA) is necessary in order to open new avenues for treatment. The aim of this study was to characterize the CD4+ T cell population and evaluate their activation and polarization status in OA joints. Fifty-five patients with end-stage knee OA (Kellgren-Lawrence grades III-IV) who underwent surgery for total knee arthroplasty (TKA) were enrolled into this study. Matched samples of synovial membrane (SM), synovial fluid (SF) and peripheral blood (PB) were analysed for CD3+ CD4+ CD8- T cell subsets [T helper type 1 (Th1), Th2, Th17, regulatory T cells] and activation status (CD25, CD69, CD45RO, CD45RA, CD62L) by flow cytometry. Subset-specific cytokines were analysed by cytometric bead array (CBA). SM and SF samples showed a distinct infiltration pattern of CD4+ T cells. In comparison to PB, a higher amount of joint-derived T cells was polarized into CD3+ CD4+ CD8- T cell subsets, with the most significant increase for proinflammatory Th1 cells in SF. CBA analysis revealed significantly increased immunomodulating cytokines [interferon (IFN)-γ, interleukin (IL)-2 and IL-10] in SF compared to PB. Whereas in PB only a small proportion of CD4+ T cells were activated, the majority of joint-derived CD4+ T cells can be characterized as activated effector memory cells (CD69+ CD45RO+ CD62L- ). End-stage OA knees are characterized by an increased CD4+ T cell polarization towards activated Th1 cells and cytokine secretion compared to PB. This local inflammation may contribute to disease aggravation and eventually perpetuate the disease process.

DNA Methyltransferase Inhibition Promotes Th1 Polarization in Human CD4+CD25high FOXP3+ Regulatory T Cells but Does Not Affect Their Suppressive Capacity.

Regulatory T cells (Treg) can show plasticity whereby FOXP3 expression, the master transcription factor for Treg suppressor function, is lost and proinflammatory cytokines are produced. Optimal FOXP3 expression strongly depends on hypomethylation of the FOXP3 gene. 5-Azacytidine (Aza) and its derivative 5-aza-2'-deoxycytidine (DAC) are DNA methyltransferase inhibitors (DNMTi) that are therapeutically used in hematological malignancies, which might be an attractive strategy to promote Treg stability. Previous in vitro research primarily focused on Treg induction by DAC from naïve conventional CD4+ T cells (Tconv). Here, we examined the in vitro effect of DAC on the stability and function of FACS-sorted human naturally occurring CD4+CD25high FOXP3+ Treg. We found that in vitro activation of Treg in the presence of DAC led to a significant inhibition of Treg proliferation, but not of Tconv. Although Treg activation in the presence of DAC led to increased IFNγ expression and induction of a Thelper-1 phenotype, the Treg maintained their suppressive capacity. DAC also induced a trend towards increased IL-10 expression. In vivo studies in patients with hematological malignancies that were treated with 5-azacytidine (Vidaza) supported the in vitro findings. In conclusion, despite its potential to increase IFNγ expression, DAC does preserve the suppressor phenotype of naturally occurring Treg.

Combination of TLR8 and TLR4 agonists reduces the degrading effects of nicotine on DC-NK mediated effector T cell generation.

The magnitude of immune responses to vaccination is a critical factor in determining protection from disease. It is known that cigarette smoke dampens the immune system and increases the risk of vaccine-preventable diseases. We reported that nicotine, the immunosuppressive component of cigarette smoke, disrupts the differentiation and functional properties of DC, which are pivotal in the initiation of immune response to vaccines. We also reported that TLR agonists act in synergy and boost DC maturation, DC-NK crosstalk and ultimately naïve T cell polarization into effector Th1 and Tc1 cells. Here, we investigated whether the combination of TLR agonists could diminish the degrading effects of nicotine on DC-NK mediated effector T cell generation. We found that none of TLR agonists, single or combined, were able to diminish completely the adverse effects of nicotine on DC. However, TLR3, TLR4, and TLR8 agonists acted as the most effective adjuvants to increase the expression levels of antigen-presenting, costimulatory molecules and production of cytokines by nicotine-exposed DC (nicDC). When combined, TLR3 + 8 and TLR4 + 8 synergistically optimized nicDC maturation and IFN-γ secretion from nicotine-exposed NK (nicNK) during co-cultures. Interestingly, in contrast to DC-NK-T, co-cultures of nicDC-nicNK-T treated with TLR3 + 8 or TLR4 + 8 agonists produced a similar frequency of effector memory Th1 and Tc1 cells. However, the effector cells from TLR4 + 8 followed by TLR3 + 8 treated nicDC-nicNK-T co-cultures produced significantly more IFN-γ when compared with aluminum salt treated co-culture. Our data suggest that addition of appropriate TLR agonists to vaccine formulation could potentially augment the immune response to vaccination in smokers.

Association of Hidradenitis Suppurativa With T Helper 1/T Helper 17 Phenotypes: A Semantic Map Analysis.

Importance: In spite of progress in understanding the mechanisms underlying hidradenitis suppurativa (HS) as an inflammatory skin disease, there is still a demand for an overview on immunopathogenesis of HS. Objective: To demonstrate the importance of the type 1/type 17 immune response in lesional HS skin by drawing a semantic connectivity map. Design, Setting, and Participants: Single-center case series of 24 patients with HS. Association of HS with T helper 1/T helper 17 (TH1/TH17) phenotype was assessed using semantic map analysis. Main Outcomes and Measures: Association of HS with TH1/TH17 phenotype. Results: The analysis was performed on 24 lesional HS biopsy samples from untreated patients with HS (16 [67%] female; median age, 36.5 years [range, 21-51 years]) with a mean (SD) Hurley stage of 2.29 (0.62) and 9 punch biopsy samples from healthy controls (6 [67%] female; median age, 43 years [range, 23-66 years]). The map shows a clustering of all TH1/TH17-associated cytokines (interleukin 17 [IL-17], interferon γ, IL-12, IL-23, IL-32, IL-1ß, tumor necrosis factor) around overall lesional inflammation. Tumor necrosis factor, IL-12, and IL-17 are even directly connected via interferon γ. In contrast, IL-13, a TH2-associated cytokine, was inversely correlated with the presence of TH1/TH17-associated cytokines, further highlighting the importance of the TH1/TH17 cytokines in HS pathogenesis. Conclusions and Relevance: These findings suggest that HS may be a TH1/TH17-driven inflammatory skin disease.

CCDC134 ameliorated experimental autoimmune encephalomyelitis by suppressing Th1 and Th17 cells.

CCDC134 (coiled-coil domain containing 134), a cytokine-like molecule, was previously reported to exert antitumor effects by augmenting CD8+ T-cell mediated immunity. However, the dynamic changes in CCDC134 expression patterns in the spinal cord that may be involved in the progression of experimental autoimmune encephalomyelitis (EAE), an animal model of multiple sclerosis, remains unclear. In this study, we found that CCDC134 expression was markedly increased in the spinal cord during the progression of EAE. Furthermore, we demonstrated that CCDC134 significantly reduced the severity and slowed the progression of EAE, which correlated with reduced spinal cord inflammation and demyelination. The underlying mechanism of CCDC134-induced effects involved inhibition of T helper (Th)-1 and Th17 cell differentiation and secretion of its key effector molecules IFN-γ and IL-17A via regulation of JAK/STAT signaling. These findings indicate that CCDC134 exerts potent anti-inflammatory effects through the selective modulation of pathogenic Th1 and Th17 cells by targeting critical signaling pathways. The study provides insights into the role of CCDC134 as a unique therapeutic agent for the treatment of autoimmune diseases.

Interleukin 35: Inhibitory regulator in monocyte-derived dendritic cell maturation and activation.

IL-35, a novel IL-12 family member, is a potent inhibitory cytokine predominantly produced by regulatory T and B lymphocytes that exerts optimal suppression in immune response. However, it remains unclear whether IL-35 plays an inhibitory role on human dendritic cells. In the present study, we focused on the possible immunosuppressive effect of IL-35 on the differentiation, maturation and function of monocyte-derived DCs (MoDCs). Addition of exogenous IL-35 was able to partially suppress MoDCs differentiation in vitro. Subsequently, LPS was used for the maturation of MoDCs and IL-35 was found to mainly restrain the maturation of MoDCs, characterized by the remarkable down-regulation of costimulatory molecules, CD83 and HLA-DR as well as a reduced production of pro-inflammatory cytokines (IL-12p70, IFN-γ, and TNF-α). Furthermore, IL-35-treated MoDCs exhibited strong inhibition in the proliferation of allogeneic CD4+/CD8+ T lymphocytes. Meanwhile, IL-35-treated MoDCs also suppressed the polarization of naïve CD4+ T lymphocytes towards Th1 phenotype and impaired CD8+ T cells allogeneic responses. And the foregoing suppression of MoDCs maturation and function by IL-35 might be due to the aberrant activation of STAT1/STAT3 and inhibition of p38 MAPK/NF-κB signaling pathway. Our results demonstrated for the first time that IL-35 played a critical role in modulating not only adaptive immune response, but also innate immune response. The inhibitory effect of IL-35 on MoDCs maturation and function may facilitate the development of promising therapeutic interventions in tumors and other diseases.

Transcriptome analysis of low-dose ionizing radiation-impacted genes in CD4+ T-cells undergoing activation and regulation of their expression of select cytokines.

Immune cells are known as the most sensitive tissue for ionizing radiation. Numerous reports relating with the effect of low-dose ionizing radiation (LDIR) on immune activities showed that LDIR can induce immune-potentiation via modulating the activity of B-, T-, and NK cells, or macrophages, whereas high-dose radiation induces genome-wide apoptotic/necrotic tissue injury and immune suppression. Generally, CD4+ T-cells play pivotal roles in immune systems via cytokines and cell-surface molecules to activate other types of immune cells to eliminate the pathogen. In spite of the significance of CD4+ T-cells in the immune system, mechanism of how LDIR regulates CD4+ T-cell gene expression is poorly investigated. Thus, RNA-Seq and Gene-Set Enrichment Analysis (GSEA) analysis were done with low-dose irradiated (γ-radiation, 50 mGy, 204 mGy/h)/anti-CD3/CD28-stimulated CD4+ T-cells to explore the LDIR-specific regulation of CD4+ T-cell gene expression. The results indicated that the genes related to mRNA translation processes, mitochondrial function, cell cycle regulation, and cytokine induction were upregulated in irradiated cells. Moreover, this study showed that the expression of T-helper cell Type 1 (TH1) or type 2 (TH2) cytokine genes, such as those for interferon (IFN)-γ, interleukin (IL)-4, and IL-5 were increased by at least 1.4-fold in acute (204 mGy/h) or chronic (10 mGy/h) low-dose (10 or 50 mGy) irradiated/anti-CD3/CD28 stimulated CD4+ T-cells, whereas the T-regulatory (Treg) cell cytokine gene, transforming growth factor (TGF)-ß was decreased. Overall, these findings demonstrated that LDIR could cause an upregulation of selected immune product genes and, in turn, might modulate the activity of CD4+ T-cells undergoing activation via an impact on cytokine gene regulation.

[MiRNAs: new actors in the physiopathology of multiple sclerosis]. / Les microARN - Nouveaux acteurs dans la physiopathologie de la sclérose en plaques.

Multiple sclerosis (MS) is an auto-immune demyelinating disorder characterized by a chronic neuro-inflammatory process associated with an infiltration of the central nervous system (CNS) by autoreactive lymphocytes. The etiology of the disease remains unclear but the recent discovery of a dysregulated miRNA network in both cells and extracellular fluids of MS patients has brought new insights on the pathophysiological mechanisms involved in this disorder. miRNAs can induce a T cell polarization towards a pathological Th17 or Th1 phenotype and a deleterious activation of microglia, the CNS-resident macrophages. We provide here a review of the most recent data regarding miRNA dysregulation and pathophysiological roles in MS patients and in the animal model of MS, EAE (experimental autoimmune encephalomyelitis). Moreover, we discuss the putative clinical value of miRNAs as a novel biomarker and diagnostic tool for MS.

TIM-4 Identifies IFN-γ-Expressing Proinflammatory B Effector 1 Cells That Promote Tumor and Allograft Rejection.

B cells give rise to polarized subsets, including B effector 1 (Be1) cells and regulatory B cells, which can promote or inhibit immune responses through expression of IFN-γ and IL-10, respectively. Such subsets likely explain why B cell depletion can either ameliorate or exacerbate inflammatory diseases; however, these cells remain poorly understood because of the absence of specific markers. Although T cell Ig and mucin domain-containing molecule (TIM)-1 broadly identifies IL-10+ regulatory B cells, no similar markers for Be1 cells have been described. We now show that TIM-4 is expressed by a subset of B cells distinct from those expressing TIM-1. Although TIM-1+ B cells are enriched for IL-10, TIM-4+ B cells are enriched for IFN-γ. TIM-1+ B cells enhanced the growth of B16-F10 melanoma. In contrast, TIM-4+ B cells decreased B16-F10 metastasis and s.c. tumor growth, and this was IFN-γ dependent. TIM-1+ B cells prolonged islet allograft survival in B-deficient mice, whereas TIM-4+ B cells accelerated rejection in an IFN-γ-dependent manner. Moreover, TIM-4+ B cells promoted proinflammatory Th differentiation in vivo, increasing IFN-γ while decreasing IL-4, IL-10, and Foxp3 expression by CD4+ T cells-effects that are opposite from those of TIM-1+ B cells. Importantly, a monoclonal anti-TIM-4 Ab promoted allograft tolerance, and this was dependent on B cell expression of TIM-4. Anti-TIM-4 downregulated T-bet and IFN-γ expression by TIM-4+ B cells and indirectly increased IL-10 expression by TIM-1+ B cells. Thus, TIM-4+ B cells are enriched for IFN-γ-producing proinflammatory Be1 cells that enhance immune responsiveness and can be specifically targeted with anti-TIM-4.

p38α regulates cytokine-induced IFNγ secretion via the Mnk1/eIF4E pathway in Th1 cells.

The p38 mitogen-activated protein kinase (MAPK) pathway is involved in the regulation of immune and inflammatory processes. We used p38α-conditional, p38ß-deficient and p38α/ß double-null mouse models to address the role of these two p38 MAPK in CD4+ T cells, and found that p38α deficiency causes these cells to hyperproliferate. Our studies indicate that both p38α and p38ß are dispensable for T helper cell type 1 (Th1) differentiation but, by controlling interferon (IFN)γ and tumor necrosis factor (TNF)α production, are critical for normal Th1 effector function. We found that both p38α and p38ß modulate T-cell receptor-induced IFNγ and TNFα production, whereas only p38α regulates cytokine-induced IFNγ production. The lack of p38α and p38ß did not affect transcription and mRNA stability of Ifng. However, the absence of p38α in Th1 cells resulted in a decreased MNK1 phosphorylation after cytokine activation, and MNK1 inhibition blocked IFNγ production. Our results indicate that p38α regulates IFNγ secretion through the activation of the MNK1/eIF4E pathway of translation initiation and identify specific functions for p38α and p38ß in T-cell proliferation.

TCR Signal Strength Regulates Akt Substrate Specificity To Induce Alternate Murine Th and T Regulatory Cell Differentiation Programs.

The Akt/mTOR pathway is a key driver of murine CD4+ T cell differentiation, and induction of regulatory T (Treg) cells results from low TCR signal strength and low Akt/mTOR signaling. However, strong TCR signals induce high Akt activity that promotes Th cell induction. Yet, it is unclear how Akt controls alternate T cell fate decisions. We find that the strength of the TCR signal results in differential Akt enzymatic activity. Surprisingly, the Akt substrate networks associated with T cell fate decisions are qualitatively different. Proteomic profiling of Akt signaling networks during Treg versus Th induction demonstrates that Akt differentially regulates RNA processing and splicing factors to drive T cell differentiation. Interestingly, heterogeneous nuclear ribonucleoprotein (hnRNP) L or hnRNP A1 are Akt substrates during Treg induction and have known roles in regulating the stability and splicing of key mRNAs that code for proteins in the canonical TCR signaling pathway, including CD3ζ and CD45. Functionally, inhibition of Akt enzymatic activity results in the dysregulation of splicing during T cell differentiation, and knockdown of hnRNP L or hnRNP A1 results in the lower induction of Treg cells. Together, this work suggests that a switch in substrate specificity coupled to the phosphorylation status of Akt may lead to alternative cell fates and demonstrates that proteins involved with alternative splicing are important factors in T cell fate decisions.

RIG-I-like Receptor Triggering by Dengue Virus Drives Dendritic Cell Immune Activation and TH1 Differentiation.

Dengue virus (DENV) causes 400 million infections annually and is one of several viruses that can cause viral hemorrhagic fever, which is characterized by uncontrolled immune activation resulting in high fever and internal bleeding. Although the underlying mechanisms are unknown, massive cytokine secretion is thought to be involved. Dendritic cells (DCs) are the main target cells of DENV, and we investigated their role in DENV-induced cytokine production and adaptive immune responses. DENV infection induced DC maturation and secretion of IL-1ß, IL-6, and TNF. Inhibition of DENV RNA replication abrogated these responses. Notably, silencing of RNA sensors RIG-I or MDA5 abrogated DC maturation, as well as cytokine responses by DENV-infected DCs. DC maturation was induced by type I IFN responses because inhibition of IFN-α/ß receptor signaling abrogated DENV-induced DC maturation. Moreover, DENV infection of DCs resulted in CCL2, CCL3, and CCL4 expression, which was abrogated after RIG-I and MDA5 silencing. DCs play an essential role in TH cell differentiation, and we show that RIG-I and MDA5 triggering by DENV leads to TH1 polarization, which is characterized by high levels of IFN-γ. Notably, cytokines IL-6, TNF, and IFN-γ and chemokines CCL2, CCL3, and CCL4 have been associated with disease severity, endothelial dysfunction, and vasodilation. Therefore, we identified RIG-I and MDA5 as critical players in innate and adaptive immune responses against DENV, and targeting these receptors has the potential to decrease hemorrhagic fever in patients.