Batf-mediated epigenetic control of effector CD8+ T cell differentiation.

The response of naive CD8+ T cells to their cognate antigen involves rapid and broad changes to gene expression that are coupled with extensive chromatin remodeling, but the mechanisms governing these changes are not fully understood. Here, we investigated how these changes depend on the basic leucine zipper ATF-like transcription factor Batf, which is essential for the early phases of the process. Through genome scale profiling, we characterized the role of Batf in chromatin organization at several levels, including the accessibility of key regulatory regions, the expression of their nearby genes, and the interactions that these regions form with each other and with key transcription factors. We identified a core network of transcription factors that cooperated with Batf, including Irf4, Runx3, and T-bet, as indicated by their colocalization with Batf and their binding in regions whose accessibility, interactions, and expression of nearby genes depend on Batf. We demonstrated the synergistic activity of this network by overexpressing the different combinations of these genes in fibroblasts. Batf and Irf4, but not Batf alone, were sufficient to increase accessibility and transcription of key loci, normally associated with T cell function. Addition of Runx3 and T-bet further contributed to fine-tuning of these changes and was essential for establishing chromatin loops characteristic of T cells. These data provide a resource for studying the epigenomic and transcriptomic landscape of effector differentiation of cytotoxic T cells and for investigating the interdependency between transcription factors and its effects on the epigenome and transcriptome of primary cells.

The cell-type specific role of Arabidopsis bZIP59 transcription factor in plant immunity.

Stomatal movement participates in plant immunity by directly affecting the invasion of bacteria, but the genes that regulate stomatal immunity have not been well identified. Here, we characterised the function of the bZIP59 transcription factor from Arabidopsis thaliana, which is constitutively expressed in guard cells. The bzip59 mutant is partially impaired in stomatal closure induced by Pseudomonas syringae pv. tomato strain (Pst) DC3000 and is more susceptible to Pst DC3000 infection. By contrast, the line overexpressing bZIP59 enhances resistance to Pst DC3000 infection. Furthermore, the bzip59 mutant is also partially impaired in stomatal closure induced by flagellin flg22 derived from Pst DC3000, and epistasis analysis revealed that bZIP59 acts upstream of reactive oxygen species (ROS) and nitric oxide (NO) and downstream of salicylic acid signalling in flg22-induced stomatal closure. In addition, the bzip59 mutant showed resistance and sensitivity to Sclerotinia sclerotiorum and Tobacco mosaic virus that do not invade through stomata, respectively. Collectively, our results demonstrate that bZIP59 plays an important role in the stomatal immunity and reveal that the same transcription factor can positively and negatively regulate disease resistance against different pathogens.

Conserved human effector Treg cell transcriptomic and epigenetic signature in arthritic joint inflammation.

Treg cells are critical regulators of immune homeostasis, and environment-driven Treg cell differentiation into effector (e)Treg cells is crucial for optimal functioning. However, human Treg cell programming in inflammation is unclear. Here, we combine transcriptional and epigenetic profiling to identify a human eTreg cell signature. Inflammation-derived functional Treg cells have a transcriptional profile characterized by upregulation of both a core Treg cell (FOXP3, CTLA4, TIGIT) and effector program (GITR, BLIMP-1, BATF). We identify a specific human eTreg cell signature that includes the vitamin D receptor (VDR) as a predicted regulator in eTreg cell differentiation. H3K27ac/H3K4me1 occupancy indicates an altered (super-)enhancer landscape, including enrichment of the VDR and BATF binding motifs. The Treg cell profile has striking overlap with tumor-infiltrating Treg cells. Our data demonstrate that human inflammation-derived Treg cells acquire a conserved and specific eTreg cell profile guided by epigenetic changes, and fine-tuned by environment-specific adaptations.

Single-cell chromatin accessibility landscape identifies tissue repair program in human regulatory T cells.

Murine regulatory T (Treg) cells in tissues promote tissue homeostasis and regeneration. We sought to identify features that characterize human Treg cells with these functions in healthy tissues. Single-cell chromatin accessibility profiles of murine and human tissue Treg cells defined a conserved, microbiota-independent tissue-repair Treg signature with a prevailing footprint of the transcription factor BATF. This signature, combined with gene expression profiling and TCR fate mapping, identified a population of tissue-like Treg cells in human peripheral blood that expressed BATF, chemokine receptor CCR8 and HLA-DR. Human BATF+CCR8+ Treg cells from normal skin and adipose tissue shared features with nonlymphoid T follicular helper-like (Tfh-like) cells, and induction of a Tfh-like differentiation program in naive human Treg cells partially recapitulated tissue Treg regenerative characteristics, including wound healing potential. Human BATF+CCR8+ Treg cells from healthy tissue share features with tumor-resident Treg cells, highlighting the importance of understanding the context-specific functions of these cells.

Adult T-cell leukemia-lymphoma as a viral disease: Subtypes based on viral aspects.

Adult T-cell leukemia-lymphoma (ATL) is caused by human T-cell leukemia virus type 1 (HTLV-1) infection. Among HTLV-1 encoded genes, HTLV-1 bZIP factor (HBZ) and tax are critical for the leukemogenesis of ATL. Adult T-cell leukemia-lymphoma needs a long latent period before onset, indicating that both viral genes and alterations (genetic and epigenetic) of the host genome play important roles for leukemogenesis. Viral genes influence genetic and epigenetic changes of the host genome, indicating that the virus is of primary importance in leukemogenesis. HBZ is expressed in all ATL cases, whereas Tax expression is heterogeneous among ATL cases. Different patterns of viral gene expression in tumors are also observed for Epstein-Barr virus. We propose three subtypes of ATL cases based on Tax expression: high, intermittent, and lost expression. HBZ is detected in all ATL cases. Approximately 25% of all ATL cases lost Tax expression at infection of HTLV-1, indicating that HBZ is the only viral gene responsible for leukemogenesis in addition to genetic and epigenetic changes of the host genes in these ATL cases. The host immune responses to Tax are also implicated in the heterogeneity of ATL. Thus, ATL is a heterogeneous disease in terms of its viral gene expression, which is important for pathogenesis of this intractable lymphomatous neoplasm.

Seroprevalence of human herpesvirus 8 in Ireland among blood donors, men who have sex with men, and heterosexual genitourinary medicine and infectious diseases clinic attendees.

Human herpesvirus 8 (HHV-8) seroprevalence varies geographically and between subpopulations. High seroprevalence rates have been ascribed to men who have sex with men (MSM), African migrants, and HIV-infected individuals. The objective of this study was to determine the seroprevalence of HHV-8 in an Irish population, including specific risk groups. A cross-sectional study of 200 blood donors and 200 genitourinary medicine (GUM) and infectious diseases (ID) clinic patients was performed, with testing for Immunoglobulin G (IgG) antibodies to HHV-8 lytic antigens using a commercial indirect fluorescence assay (Scimedx Corp.). Verification was performed at the Centers for Disease Control and Prevention (CDC). All 200 blood donor samples were negative for HHV-8 IgG antibodies. 21% of GUM and ID patients were positive for HHV-8 IgG antibodies. One hundred of these patients were MSM, 35% of whom were HHV-8 seropositive (46% of HIV-positive MSM and 24% of HIV-negative MSM). Of 100 heterosexual patients, only 7% were HHV-8 seropositive. The absence of seropositivity in 200 Irish blood donors may suggest that Ireland has a low overall population HHV-8 seroprevalence. The proportion of HHV-8 seropositivity in the MSM population was significantly higher than in the heterosexual population and most marked in HIV-positive MSM.

IL-4-BATF signaling directly modulates IL-9 producing mucosal mast cell (MMC9) function in experimental food allergy.

BACKGROUND: This study group has previously identified IL-9-producing mucosal mast cell (MMC9) as the primary source of IL-9 to drive intestinal mastocytosis and experimental IgE-mediated food allergy. However, the molecular mechanisms that regulate the expansion of MMC9s remain unknown. OBJECTIVES: This study hypothesized that IL-4 regulates MMC9 development and MMC9-dependent experimental IgE-mediated food allergy. METHODS: An epicutaneous sensitization model was used and bone marrow reconstitution experiments were performed to test the requirement of IL-4 receptor α (IL-4Rα) signaling on MMC9s in experimental IgE-mediated food allergy. Flow cytometric, bulk, and single-cell RNA-sequencing analyses on small intestine (SI) MMC9s were performed to illuminate MMC9 transcriptional signature and the effect of IL-4Rα signaling on MMC9 function. A bone marrow-derived MMC9 culture system was used to define IL-4-BATF signaling in MMC9 development. RESULTS: Epicutaneous sensitization- and bone marrow reconstitution-based models of IgE-mediated food allergy revealed an IL-4 signaling-dependent cell-intrinsic effect on SI MMC9 accumulation and food allergy severity. RNA-sequencing analysis of SI-MMC9s identified 410 gene transcripts reciprocally regulated by IL-4 signaling, including Il9 and Batf. Insilico analyses identified a 3491-gene MMC9 transcriptional signature and identified 2 transcriptionally distinct SI MMC9 populations enriched for metabolic or inflammatory programs. Employing an in vitro MMC9-culture model system showed that generation of MMC9-like cells was induced by IL-4 and this was in part dependent on BATF. CONCLUSIONS: IL-4Rα signaling directly modulates MMC9 function and exacerbation of experimental IgE-mediated food allergic reactions. IL-4Rα regulation of MMC9s is in part BATF-dependent and occurs via modulation of metabolic transcriptional programs.

Resistance to Experimental Visceral Leishmaniasis in Mice Infected With Leishmania infantum Requires Batf3.

Unveiling the protective immune response to visceral leishmaniasis is critical for a rational design of vaccines aimed at reducing the impact caused by this fatal, if left untreated, vector-borne disease. In this study we sought to determine the role of the basic leucine zipper transcription factor ATF-like 3 (Batf3) in the evolution of infection with Leishmania infantum, the causative agent of human visceral leishmaniasis in the Mediterranean Basin and Latin America. For that, Batf3-deficient mice in C57BL/6 background were infected with an L. infantum strain expressing the luciferase gene. Bioluminescent imaging, as well as in vitro parasite titration, demonstrated that Batf3-deficient mice were unable to control hepatic parasitosis as opposed to wild-type C57BL/6 mice. The impaired microbicide capacities of L. infantum-infected macrophages from Batf3-deficient mice mainly correlated with a reduction of parasite-specific IFN-γ production. Our results reinforce the implication of Batf3 in the generation of type 1 immunity against infectious diseases.

CD103+ cDC1 and endogenous CD8+ T cells are necessary for improved CD40L-overexpressing CAR T cell antitumor function.

While effective in specific settings, adoptive chimeric antigen receptor (CAR) T cell therapy for cancer requires further improvement and optimization. Our previous results show that CD40L-overexpressing CAR T cells mobilize endogenous immune effectors, resulting in improved antitumor immunity. However, the cell populations required for this protective effect remain to be identified. Here we show, by analyzing Batf3-/- mice lacking the CD103+ conventional dendritic cell type 1 (cDC1) subpopulation important for antigen cross-presentation, that CD40L-overexpressing CAR T cells elicit an impaired antitumor response in the absence of cDC1s. We further find that CD40L-overexpressing CAR T cells stimulate tumor-resident CD11b-CD103- double-negative (DN) cDCs to proliferate and differentiate into cDC1s in wild-type mice. Finally, re-challenge experiments show that endogenous CD8+ T cells are required for protective antitumor memory in this setting. Our findings thus demonstrate the stimulatory effect of CD40L-overexpressing CAR T cells on innate and adaptive immune cells, and provide a rationale for using CD40L-overexpressing CAR T cells to improve immunotherapy responses.

STAT5 promotes accessibility and is required for BATF-mediated plasticity at the Il9 locus.

T helper cell differentiation requires lineage-defining transcription factors and factors that have shared expression among multiple subsets. BATF is required for development of multiple Th subsets but functions in a lineage-specific manner. BATF is required for IL-9 production in Th9 cells but in contrast to its function as a pioneer factor in Th17 cells, BATF is neither sufficient nor required for accessibility at the Il9 locus. Here we show that STAT5 is the earliest factor binding and remodeling the Il9 locus to allow BATF binding in both mouse and human Th9 cultures. The ability of STAT5 to mediate accessibility for BATF is observed in other Th lineages and allows acquisition of the IL-9-secreting phenotype. STAT5 and BATF convert Th17 cells into cells that mediate IL-9-dependent effects in allergic airway inflammation and anti-tumor immunity. Thus, BATF requires the STAT5 signal to mediate plasticity at the Il9 locus.

Cutting Edge: Batf3 Expression by CD8 T Cells Critically Regulates the Development of Memory Populations.

The basic leucine zipper transcription factor ATF-like 3 (BATF3) is required for the development of conventional type 1 dendritic cells that are essential for cross-presentation and CD8 T cell-mediated immunity against intracellular pathogens and tumors. However, whether BATF3 intrinsically regulates CD8 T cell responses is not well studied. In this article, we report a role for cell-intrinsic Batf3 expression in regulating the establishment of circulating and resident memory T cells after foodborne Listeria monocytogenes infection of mice. Consistent with other studies, Batf3 expression by CD8 T cells was dispensable for the primary response. However, Batf3 -/- T cells underwent increased apoptosis during contraction to contribute to a substantially reduced memory population. Batf3 -/- memory cells had an impaired ability to mount a robust recall response but remained functional. These findings reveal a cell-intrinsic role of Batf3 in regulating CD8 T cell memory development.

HTLV-1 induces T cell malignancy and inflammation by viral antisense factor-mediated modulation of the cytokine signaling.

Human T cell leukemia virus type 1 (HTLV-1) is the etiologic agent of a T cell neoplasm and several inflammatory diseases. A viral gene, HTLV-1 bZIP factor (HBZ), induces pathogenic Foxp3-expressing T cells and triggers systemic inflammation and T cell lymphoma in transgenic mice, indicating its significance in HTLV-1-associated diseases. Here we show that, unexpectedly, a proinflammatory cytokine, IL-6, counteracts HBZ-mediated pathogenesis. Loss of IL-6 accelerates inflammation and lymphomagenesis in HBZ transgenic mice. IL-6 innately inhibits regulatory T cell differentiation, suggesting that IL-6 functions as a suppressor against HBZ-associated complications. HBZ up-regulates expression of the immunosuppressive cytokine IL-10. IL-10 promotes T cell proliferation only in the presence of HBZ. As a mechanism of growth promotion by IL-10, HBZ interacts with STAT1 and STAT3 and modulates the IL-10/JAK/STAT signaling pathway. These findings suggest that HTLV-1 promotes the proliferation of infected T cells by hijacking the machinery of regulatory T cell differentiation. IL-10 induced by HBZ likely suppresses the host immune response and concurrently promotes the proliferation of HTLV-1 infected T cells.

The IL-18, IL-12, and IFN-γ expression in HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP) patients, HTLV-1 carriers, and healthy subjects.

Interleukin (IL)-12, IL-18, and interferon gamma (IFN-γ) can induce Th1-inflammatory responses in favor of HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP) manifestation. In this study, the gene expression and plasma levels of these cytokines were evaluated. The peripheral blood mononuclear cells (PBMCs) in 20 HAM/TSP patients, 21 asymptomatic carriers (ACs), and 21 healthy subjects (HSs) were assessed for the expression of IL-18, IL-12, and IFN-γ, using qRT-PCR. The plasma level of IL-18 and IFN-γ were measured by an ELISA method. The mean of HTLV-1 proviral load (PVL) in the HAM/TSPs was 1846.59 ± 273.25 and higher than ACs at 719.58 ± 150.72 (p = 0.001). The IL-12 was considerably expressed only in nine ACs, five HAM/TSPs, and all HSs. Furthermore, the gene expression and plasma levels of IL-18 were lower in the HTLV-1-positive group than the control group (p = 0.001 and 0.012, respectively); however, there was no significant difference between the ACs and HAM/TSPs. The IFN-γ level was higher in the HTLV-1-positive group (p < 0.001) than HSs. Although there were no correlation between plasma levels of IL-18 and IFN-γ with PVL in the ACs, a positive correlation was observed between plasma IL-18 levels and PVL (r = 0.654, p = 0.002). The highest levels of IFN-γ were observed in the HAM/TSPs which has a significant correlation with HTLV-1-HBZ (r = 0.387, p = 0.05) but not with Tax. However, no significant correlation was found between PVL and proinflammatory pattern. Apart from the IFN-γ as a lymphokine, as a host factor, and HTLV-1-HBZ, as a viral agent, the other proinflammatory monokines or HTLV-1 factors are among the less-effective agents in the maintenance of HAM/TSP.

Expression of TSLC1 in patients with HAM/TSP.

Human T-lymphotropic virus type 1 (HTLV-1)-associated myelopathy/tropical spastic paraparesis (HAM/TSP) is chronic myelopathy characterized by slowly progressive spastic paraparesis and urinary dysfunction. A few biomarkers in the cerebrospinal fluid are known to be related to disease activity, but no biomarker has been reported in peripheral blood. This study aims to explore the expression level of the adhesion molecule during the expression level of the adhesion molecule among HAM/TSP disease activity. In lymphocyte function-associated antigen 1 and DNAX accessory molecule 1, no variation in expression levels specific to HTLV-1 infection was observed in CD4-positive T cells; however, TSLC1 expression was higher in HAM patients than in asymptomatic carriers and non-infected persons. TSLC1 tended to be higher in patients whose symptoms were worsening. On the contrary, the expression level of TSLC1 in CD8-positive T cells was lower in HAM patients than in asymptomatic carriers, and this tendency was stronger in patients whose symptoms had deteriorated. No significant correlation was found between TSLC1 and either of the transcription factors Tax or HBZ in any T cell group. Therefore, TSLC1 expression in CD4-positive T cells might be a useful biomarker of HAM/TSP disease activity.

E4BP4-mediated inhibition of T follicular helper cell differentiation is compromised in autoimmune diseases.

T follicular helper (Tfh) cells are indispensable for the formation of germinal center (GC) reactions, whereas T follicular regulatory (Tfr) cells inhibit Tfh-mediated GC responses. Aberrant activation of Tfh cells contributes substantially to the pathogenesis of autoimmune diseases, such as systemic lupus erythematosus (SLE). Nonetheless, the molecular mechanisms mitigating excessive Tfh cell differentiation are not fully understood. Herein we demonstrate that the adenovirus E4 promoter-binding protein (E4BP4) mediates a feedback loop and acts as a transcriptional brake to inhibit Tfh cell differentiation. Furthermore, we show that such an immunological mechanism is compromised in patients with SLE. Establishing mice with either conditional knockout (cKO) or knockin (cKI) of the E4bp4 gene in T cells reveals that E4BP4 strongly inhibits Tfh cell differentiation. Mechanistically, E4BP4 regulates Bcl6 transcription by recruiting the repressive epigenetic modifiers HDAC1 and EZH2. E4BP4 phosphorylation site mutants have limited capability with regard to inhibiting Tfh cell differentiation. In SLE, we detected impaired phosphorylation of E4BP4, finding that this compromised transcription factor is positively correlated with disease activity. These findings unveiled molecular mechanisms by which E4BP4 restrains Tfh cell differentiation, whose compromised function is associated with uncontrolled autoimmune reactions in SLE.

Type I Interferon Response Dysregulates Host Iron Homeostasis and Enhances Candida glabrata Infection.

Type I interferons (IFNs-I) fulfil multiple protective functions during pathogenic infections, but they can also cause detrimental effects and enhance immunopathology. Here, we report that IFNs-I promote the dysregulation of iron homeostasis in macrophages during systemic infections with the intracellular pathogen Candida glabrata, leading to fungal survival and persistence. By engaging JAK1, IFNs-I disturb the balance of the transcriptional activator NRF2 and repressor BACH1 to induce downregulation of the key iron exporter Fpn1 in macrophages. This leads to enhanced iron accumulation in the phagolysosome and failure to restrict fungal access to iron pools. As a result, C. glabrata acquires iron via the Sit1/Ftr1 iron transporter system, facilitating fungal intracellular replication and immune evasion. Thus, IFNs-I are central regulators of iron homeostasis, which can impact infection, and restricting iron bioavailability may offer therapeutic strategies to combat invasive fungal infections.

Bach2 Deficiency Leads to Spontaneous Expansion of IL-4-Producing T Follicular Helper Cells and Autoimmunity.

The transcription factor Bach2 is a susceptible gene for numerous autoimmune diseases including systemic lupus erythematosus (SLE). Bach2-/- mice can develop a lupus-like autoimmune disease. However, the exact cellular and molecular mechanisms via which Bach2 protects the hosts from developing autoimmunity remains incompletely understood. Here, we report that Bach2 ablation on T cells, but not B cells, resulted in humoral autoimmunity, and this was associated with expansion of T follicular helper (Tfh) cells and abnormal germinal centers. Bach2 was down-regulated in Tfh cells and directly suppressed by the Tfh-defining transcription factor BCL6. Mechanistically, Bach2 directly suppresses the transcription of Cxcr5 and c-Maf, two key regulators of Tfh cell differentiation. Bach2-deficient Tfh cells were skewed toward the IL-4-producing subset, which induced IgG1 and IgE isotype switching of B cells. Heterozygous Bcl6 deficiency reduced the formation of germinal center and autoantibodies, and ameliorated the pathology in Bach2-deficient mice. Our findings identify Bach2 as a crucial negative regulator of Tfh cells at steady state and prove that Bach2 controls autoimmunity in part by restraining accumulation of pathogenic Tfh cells.

BATF3-dependent dendritic cells drive both effector and regulatory T-cell responses in bacterially infected tissues.

The gastric lamina propria of mice that have been experimentally infected with the pathobiont Helicobacter pylori hosts a dense network of myeloid cells that includes BATF3-dependent CD103+ dendritic cells (DCs). We show here that CD103+ DCs are strictly required for gastric Th1 responses to H. pylori and for H. pylori infection control. A similar dependence of type 1 immunity on CD103+ DCs is observed in a Mycobacterium bovis BCG infection model, and in a syngeneic colon cancer model. Strikingly, we find that not only the expansion and/or recruitment of Th1 cells, but also of peripherally induced, neuropilin-negative regulatory T-cells to sites of infection requires BATF3-dependent DCs. A shared feature of the examined models is the strongly reduced production of the chemokines and CXCR3 ligands CXCL9, 10 and 11 in BATF3-deficient mice. The results implicate BATF3-dependent DCs in the recruitment of CXCR3+ effector and regulatory T-cells to target tissues and in their local expansion.

Bach2 Negatively Regulates T Follicular Helper Cell Differentiation and Is Critical for CD4+ T Cell Memory.

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.

TET enzymes augment activation-induced deaminase (AID) expression via 5-hydroxymethylcytosine modifications at the Aicda superenhancer.

TET enzymes are dioxygenases that promote DNA demethylation by oxidizing the methyl group of 5-methylcytosine to 5-hydroxymethylcytosine (5hmC). Here, we report a close correspondence between 5hmC-marked regions, chromatin accessibility and enhancer activity in B cells, and a strong enrichment for consensus binding motifs for basic region-leucine zipper (bZIP) transcription factors at TET-responsive genomic regions. Functionally, Tet2 and Tet3 regulate class switch recombination (CSR) in murine B cells by enhancing expression of Aicda, which encodes the activation-induced cytidine deaminase (AID) enzyme essential for CSR. TET enzymes deposit 5hmC, facilitate DNA demethylation, and maintain chromatin accessibility at two TET-responsive enhancer elements, TetE1 and TetE2, located within a superenhancer in the Aicda locus. Our data identify the bZIP transcription factor, ATF-like (BATF) as a key transcription factor involved in TET-dependent Aicda expression. 5hmC is not deposited at TetE1 in activated Batf-deficient B cells, indicating that BATF facilitates TET recruitment to this Aicda enhancer. Our study emphasizes the importance of TET enzymes for bolstering AID expression and highlights 5hmC as an epigenetic mark that captures enhancer dynamics during cell activation.