Immunity

Viral pneumonia is perpetuated by inflammatory circuits between activated T cells and monocyte-derived alveolar macrophages (MoAM). T cells and macrophages express ORAI1 and STIM1, which form calcium release-activated calcium (CRAC) channels that allow extracellular calcium entry in response to endoplasmic reticulum calcium store depletion. In a randomized, placebo-controlled, multicenter phase 2 trial (CARDEA), Auxora, a CRAC channel inhibitor, reduced all-cause 30-day mortality by 56% in patients with severe SARS-CoV-2 pneumonia. Here, we report a multi-omics analysis of serially collected alveolar samples from unvaccinated patients with severe SARS-CoV-2 pneumonia treated with Auxora versus placebo. We found reductions in plasma levels of the monocyte- and T cell-chemokines, CCL8 and PDGF-AA. Using peripheral blood mononuclear cells (PBMC) from healthy volunteers, we show that Auxora directly targets T cells to inhibit the transcription of CCL8 and PDGFA in monocyte-derived macrophages, supporting a mechanism for its effects and a potential intermediate biomarker of efficacy.

Targeting Tregs is a potential strategy to improve cancer therapies. However, which Tregs accumulate in response to tumoral processes, and how tumors affect their phenotype, is poorly understood. Here we show that tumor Tregs are equivalent to effector tissue Tregs in steady state organs. We used a mouse model of intestinal neoplasia to demonstrate that one early event in carcinogenesis is sufficient to induce local accumulation of Tregs resembling human tumor Tregs. Treg accumulation was driven by TCR-dependent oligoclonal expansion of tissue Tregs with an effector Treg phenotype. Treg expansion was independent of CCR8, IL33R and CD137, which were previously linked to tumor Treg. In contrast, GATA3 was required for effector tissue Tregs and for their expansion in response to neoplasia. Our findings identify GATA3-dependent clonal expansion of effector tissue Tregs as a key event in promoting tumor growth. HighlightsO_LIAn early tumorigenic event alone drives accumulation of effector tissue Tregs C_LIO_LITregs in tumors are phenotypically akin to effector tissue Tregs C_LIO_LIThe accumulation of Tregs is driven by TCR-dependent oligoclonal expansion C_LIO_LIGATA3 controls tumor-promoting effector tissue Tregs C_LI

Immune responses to infection and vaccination exhibit diversity between individuals that can be shaped by differences in their immune cell landscapes and the signalling, transcriptional, and genetic mechanisms that coordinate immune cell function. Specific antibody deficiency (SAD) and common variable immunodeficiency (CVID) are common forms of predominantly antibody deficiencies that result in poor responses to vaccination. While molecular and cellular causes of the immune dysfunction and poor vaccination responses for individuals with CVID have been reported, immune cell or molecular defects have not yet been identified in SAD. Here, we have used single-cell multi-omics to define the cellular landscapes, transcriptional states, adaptive immune repertoires and protein expression of patients with SAD and CVID before and after polysaccharide vaccination. We discovered that while SAD and CVID exhibit overlapping immune defects, including accumulation of exhausted NK memory cells and dysregulated expression of genes that mediate lipopolysaccharide sensing and clearance by monocytes, individuals with SAD have a unique expansion of cytotoxic CD4+ T cells that correlates with reduced regulatory T cells. In response to vaccination, we observed rapid changes in gene expression associated with lipopolysaccharide responses by monocytes and NF-kB pathway activation in B cells, and an apparent expansion of a CD95+ class-switched memory B cell population that does not occur in patients with lower antigen-specific responses. Together, our findings reveal cellular and molecular factors that underpin variability in vaccine responses and define SAD in a broader spectrum of immune dysfunction.

Systemic Lupus Erythematosus (SLE) is a chronic autoimmune disease driven by uncensored B and T cell autoreactivity. Understanding this pathogenic process has been hampered by lack of studies of secondary lymphoid organs in human SLE. Using minimally invasive lymph node fine needle aspirates (LN-FNAs), we profiled tissue-resident immune cells from 59 SLE patients and 34 healthy controls through high-dimensional 43-color flow cytometry, antigen-specific tetramer probing, and sc-RNA sequencing with paired VH/VL repertoire analysis. Our findings reveal hyperactive lymph node immunity in SLE characterized by spontaneous germinal center (GC) activation, plasma cell accumulation enriched in mature CD19- and CD138+ antibody-secreting cells, and increased frequencies of both GC-TFH and PD-1+CXCR5- T extra-follicular helper cells. SLE lymph nodes harbored large oligoclonal B cell families with altered isotype usage, dominated by IgG1 and IgG4. Critically, self-reactive 9G4+ and Ro60+ B cells showed defective tolerance checkpoint control, accumulating in activated naive, GC, and plasma cell compartments with distinctive PD-1+Tox+ expression absent in viral-specific responses. Single-cell repertoire analysis revealed VH4-34 clones in SLE BGC and BPC, that in contrast to HD, had not experienced clonal redemption. Instead, SLE VH4-34 clones displayed low somatic hypermutation and preserved the AVY hydrophobic patch associated with autoreactivity. Monoclonal antibody testing confirmed that unmutated AVY+ VH4-34 clones retained polyreactivity against naive B cells, apoptotic cells, and multiple self-antigens. Together, these results define "clonal damnation" as a key mechanism in SLE whereby autoreactive VH4-34 clones of pathogenic potential escape tolerance checkpoints, expand in germinal centers, and differentiate into tissue plasma cells while preserving germline-encoded self-reactivity. Combined, our study defines critical mechanisms of tolerance breakdown in lupus pathogenesis.

T-cells are central to SARS-CoV-2 clearance and immunological memory, yet their contribution to the persistence of post-acute sequelae of COVID-19 (PASC) remains poorly understood. The immunological features that distinguish individuals who develop PASC from those who recover fully are unresolved, in part due to the phenotypic heterogeneity of the condition and the likely multiplicity of its underlying mechanisms. Here, we profiled longitudinal bulk TCR{beta} repertoires from 120 individuals in the INCOV cohort-71 with PASC and 49 without-sampled at two to three time points spanning the acute and post-acute phases of infection. Using robust statistical modeling of repertoire composition and clonal dynamics, we found that global statistics such as V, J gene usage and CDR3 length do not differ between groups, but that locally enriched sequence motifs and differentially dynamic clones reveal distinct T-cell signatures associated with PASC status. Clones contracting following the peak of the acute response were significantly enriched for SARS-CoV-2 specificity in both groups. Interestingly, Influenza A-specific TCRs were disproportionately enriched among contracting clones in PASC+ repertoires, implicating viral co-infection as a potential contributor to early disease severity and, possibly, PASC pathogenesis. Rare public TCR clones were markedly enriched for SARS-CoV-2 specificity, with PASC+ individuals harboring a modestly but significantly higher proportion than PASC- individuals. Together, we identified over 1,000 candidate TCR{beta} receptors potentially discriminating PASC+ from PASC- immune responses, opening a path toward the identification of disease-relevant T-cell specificities and the development of T-cell-based immunological biomarkers for long COVID.

Self-reactive B cells are generated during normal development and can acquire increased pathogenicity through activation-induced cytidine deaminase (AID)-mediated diversification following activation. Clonal deletion is thought to eliminate these cells, yet how deletion is distributed across developmental and activation stages to prevent autoimmune disease remains unclear. Here, we show that clonal deletion is enforced through temporally distinct mitochondrial apoptosis (MOMP) checkpoints that differentially regulate autoreactive B cell fate and disease progression. Using conditional Bcl-2 expression to inhibit MOMP either before or after B cell activation, we find that early inhibition permits the survival and maturation of autoreactive B cells after peripheral egress, expanding the pool of cells available for activation. These cells subsequently undergo AID-dependent diversification, producing class-switched IgG autoantibodies with expanded antigen breadth that target a wider range of self-antigens and drive lethal, female-biased autoimmune disease characterized by complement activation and kidney pathology. In contrast, inhibition of MOMP only after activation allows the accumulation of germinal center, switched memory, and plasma cells and promotes autoantibody production, but results in more restricted IgG autoreactivity, limited complement activation and limited tissue damage, and normal survival. Notably, early MOMP inhibition does not expand immature bone marrow B cells, indicating that a major clonal deletion checkpoint operates in the periphery rather than during initial B cell generation. Together, these findings support a Distributed Clonal Deletion Model in which early checkpoints restrict the entry of autoreactive B cells into diversification pathways, while later checkpoints limit the persistence of diversified autoreactive clones, thereby constraining autoimmune disease progression. One Sentence SummaryDistributed clonal deletion prevents autoimmune disease progression by restricting the breadth of autoreactive clones entering immune responses, with early MOMP checkpoints limiting diversification and later checkpoints constraining persistence.

TNF is a potent proinflammatory cytokine that can induce cell death by activating the kinase RIPK1. The adaptor proteins TANK and AZI2 protect against cell death by recruiting TBK1 to the TNF receptor signaling complex, thereby inhibiting RIPK1. While deficiency of either adaptor alone is well tolerated, combined loss of TANK and AZI2 results in partial embryonic lethality and severe TNF- and RIPK1-driven autoinflammation. Here, we show that TANK/AZI2-deficient mice exhibit a striking expansion of regulatory T cells (Tregs), most of which display an effector phenotype with high expression of immunosuppressive genes. Although thymic Treg generation is modestly increased, Tregs arise predominantly in the periphery through a largely cell-intrinsic mechanism. The marked accumulation of effector Tregs suggested that the T-cell compartment may limit TNF-driven pathology in this model. Supporting this, T cell ablation in TANK/AZI2-deficient mice markedly exacerbates disease progression and enhances TNF-driven, RIPK1-mediated inflammation. Similarly, T cells protect against acute TNF-induced systemic inflammatory response syndrome by limiting RIPK1-mediated cell death. Together, our findings identify TANK and AZI2 as negative regulators of Treg formation, demonstrate that T cells restrain TNF-driven inflammation by limiting RIPK1-dependent cell death, and suggest that this protective effect is mediated primarily by Tregs.

Variants in TNFAIP3 (encoding A20) are among the most consistent non-MHC genetic associations with human autoimmune disease. Moreover, TNFAIP3 haploinsufficiency confers an autosomal dominant autoinflammatory and autoimmune disease. Whilst these findings strongly suggest that quantitative changes in A20 predispose to pathology, there is currently no evidence that post-transcriptional modifications regulate A20 abundance. We used human reverse genetics approach to investigate this question. From a cohort of patients with complex immune diseases, we identified an ultrarare hypomorphic TNFAIP3 variant (A20S254R) that is both hypomorphic and prone to phosphorylation. One of five carriers of this variant was distinguished by autoimmunity and an NF-kB gain-of-function transcriptional signature. A search for modifiers identified a variant in TAX1BP1 (p.Leu207Ile). We discovered that TAX1BP1 normally retrains A20 phosphorylation. Furthermore, TAX1BP1 preferentially binds phospho-A20, which explains the enhanced interaction between TAX1BP1L207I and A20S254R. A20 phosphorylation regulates its abundance, not by MALT1-mediated degradation, but probably via autophagy. Thus, while A20 phosphorylation has been implicated in A20 protease activity, an epistatic interaction between rare human genetic variants reveals how phosphorylation also regulates A20 abundance.

Blocking the programmed cell death 1 (PD-1) pathway using monoclonal antibodies reinvigorates exhausted T cells (Tex), enhancing control of chronic viral infections and cancer. Considerable effort has focused on evaluating different PD-1 blockade agents in preclinical and clinical cancer settings, but relatively little information exists on how to optimize the pharmacodynamic effects of PD-1 pathway blockade on reinvigorating Tex. To address this question, we performed longitudinal tracking of Tex reinvigoration during chronic infection with lymphocytic choriomeningitis virus (LCMV) following different regimens of PD-1 blockade. We compared single-cycle (2 weeks of treatment), long-term continuous PD-1 pathway blockade (i.e. 3 months), or blockade followed by a drug holiday and then re-blockade (intermittent treatment). These studies revealed little benefit of continuous versus single-cycle PD-1 blockade, with both resulting in a single peak of Tex reinvigoration and similar effects on viral replication. In contrast, intermittent blockade resulted in a new cycle of secondary Tex reinvigoration upon redosing after a washout and this secondary Tex reinvigoration improved disease control. Mechanistically, long-term blockade eroded the ability of Tex progenitor cells (Tpex) to give rise to downstream, more functional Tex intermediate (Tex-Int) progeny, whereas the drug holiday restored this Tpex proliferative and differentiation capacity. Tpex from long-term treated mice showed evidence of adaptive resistance and additional layers of negative regulation, including sustained expression of the inhibitory receptor CD22. Indeed, co-blockade of PD-1 and CD22 using combination antibodies or bispecific antibody approaches improved disease control and reinvigoration of Tex. These data have implications for clinical immune pharmacodynamics of PD-1 blockade and provide insights into the biology of Tex reinvigoration. One Sentence SummaryModifying the immunopharmacology of PD-1 blockade reveals a benefit of a drug holiday and identifies mechanisms of Tex progenitor deficiency provoked by prolonged loss of PD-1 signals including the inhibitory receptor CD22.

Pulmonary hemorrhage (PH) is a life-threatening manifestation of systemic autoimmunity characterized by immune-mediated disruption of the alveolar-capillary barrier. Despite mortality rates exceeding 50%, the molecular checkpoints that govern the transition from destructive inflammation to protective immune regulation remain poorly defined. Building on our discovery that interleukin-2-inducible T cell kinase (ITK) uncouples pathogenic inflammation from protective immunity, we investigated ITK as a central regulator of autoimmune lung injury. Using the pristane-induced PH model, we show that ITK deficiency confers near-complete protection against PH and associated multi-organ injury. This protection is accompanied by marked remodeling of the T cell compartment, including expansion of CD44CD122EomesT-bet memory-like subsets and significant enrichment of Foxp3 regulatory T cells (Tregs). Notably, adoptive transfer of ITK-deficient Tregs was sufficient to rescue PH and suppress systemic proinflammatory cytokine production in wild-type recipients, identifying these cells as key mediators of tissue protection. Transcriptomic profiling further revealed that loss of ITK signaling reprograms Tregs toward a metabolically and functionally enhanced state, with enrichment of oxidative phosphorylation (OXPHOS), mTORC1, STAT5 signaling, and tissue-repair-associated programs. Together, these findings identify ITK as a critical regulator of the balance between pulmonary injury and reparative immunity and provide a mechanistic rationale for targeting the ITK axis in severe inflammatory lung disease. HighlightsO_LIITK deficiency protects against pristane-induced pulmonary hemorrhage (PH) C_LIO_LILoss of ITK expands "super-fit" canonical and non-canonical Tregs C_LIO_LIITK-deficient Tregs exhibit enriched mTORC1, OXPHOS, and IL-10 production C_LIO_LITransfer of ITK-deficient Tregs rescues established PH and reverses proteinuria C_LIO_LIITK uncouples pathogenic inflammation from reparative tissue immunity C_LI

Tissue-specific peripheral tolerance mechanisms are essential to prevent autoimmunity. The cornea is immune privileged, and anterior chamber-associated immune deviation (ACAID) governs its inner surface. However, the mechanisms that apply to corneal epithelial (outer surface) antigens remain unknown. Using an inducible, cornea-restricted neoantigen mouse model, we found that the cornea relies on inducible regulatory T cells (Tregs) rather than ignorance or ACAID for its epithelial antigens. Although the cornea is both avascular and alymphatic, its epithelial antigens are still efficiently presented by ocular surface-derived antigen-presenting cells to T cells in draining lymph nodes under homeostatic conditions, leading to conventional antigen-specific Treg expansion without ocular pathology. This tolerance was not absolute: systemic immunization redirected antigen-specific responses toward pathogenic effector T cells that disrupted epithelial barrier function. These findings identify Treg induction as a dominant mechanism of corneal epithelial immune homeostasis and demonstrate that inflammatory priming can render a tolerated corneal antigen into an autoimmune target, providing mechanistic insight into dry eye pathogenesis. SummaryThis study shows that immune tolerance to corneal epithelial neoantigens relies not on immune privilege but on peripherally induced regulatory T cells in the draining lymph nodes that can be subverted by innate activation, shedding light on ocular surface disease pathophysiology.

Ulcerative colitis (UC) is characterized by epithelial barrier dysfunction and dysregulated mucosal immune responses; however, the mechanisms driving disease onset remain poorly defined. Autoantibodies against the epithelial-restricted integrin v{beta}6 are a highly specific biomarker of UC that can precede clinical diagnosis by up to 10 years. Because v{beta}6 activates TGF{beta} at epithelial surfaces, we hypothesized that UC-associated v{beta}6 autoantibodies inhibit mucosal TGF{beta} activation and disrupt epithelial homeostasis. We showed that v{beta}6 autoantibodies were enriched in UC and that IgG from autoantibody-positive individuals inhibited v{beta}6-dependent activation of TGF{beta}. v{beta}6 blockade dampened TGF{beta} signaling and altered differentiation-associated gene programs in human intestinal epithelial cells. In mice, deletion of v caused expansion of inflammation-associated goblet cells in the colon and changes in intestinal immune cells. Using a novel mouse model, we showed that v{beta}6-specific autoantibody disrupted epithelial-immune crosstalk and increased susceptibility to DSS colitis. Together, these findings establish anti-v{beta}6 autoantibodies as active inhibitors of epithelial TGF{beta} signaling, constituting a de facto anti-cytokine response, rather than passive biomarkers. By linking preclinical seropositivity to impaired epithelial signaling and heightened susceptibility to colitis, this work identifies epithelial v{beta}6-dependent TGF{beta} activation as a pathway that may be leveraged to modify disease risk or limit disease severity. One Sentence SummaryUC-associated autoantibodies impair epithelial TGF{beta} activation, alter mucosal homeostasis, and predispose to colitis.

The transcription coregulator OCA-B promotes CD4+ T cell memory recall responses and autoimmunity. OCA-B T cell deletion prevents spontaneous type-1 diabetes (T1D) onset in non-obese diabetic (NOD) mice and blunts T1D in a subset of more aggressive models. However, the role of OCA-B in diabetes induced by treatment with immune checkpoint inhibitors (ICIs), and the role of OCA-B in the control of tumors with and without ICI treatment, has not been studied. Here we show that islet and pancreatic lymph node T cells from T1D individuals express measurable POU2AF1 mRNA. Deletion of OCA-B in T cells fully insulates 8-week-old non-obese diabetic (NOD) mice against ICI-induced diabetes and partially protects 12-week-old mice. Salivary and lacrimal gland infiltration and inflammation were also reduced. Protection was associated with a block in the differentiation of progenitor exhausted CD8+ T cells (TPEX) into terminally exhausted CD8+ T cells (TEX). We show that OCA-B T cell loss preserves anti-tumor immune responses following PD-1 blockade in different tumors and mouse strains. These findings point to a potential therapeutic window in which pharmaceuticals targeting OCA-B could be used to block the emergence of both spontaneous and ICI-induced autoimmunity while sparing anti-tumor immunity. We develop first-in-class small molecule inhibitors of Oct1/OCA-B transcription complexes and show that administration into NOD mice also blocks diabetes emergence following PD-1 blockade. These results identify OCA-B as a promising therapeutic target for the prevention of autoimmunity and immune-related adverse events (irAEs).

Zika virus (ZIKV) encephalitis induces cytokine-mediated cognitive deficits which persist long-term. Here, we determined if NOD2-mediated conversion of monocytes from Ly6CHi inflammatory to Ly6CLo anti-inflammatory phenotypes during ZIKV encephalitis preserves neural correlates of learning and memory within the hippocampus. Short-term administration of the NOD2 agonist, muramyl dipeptide (MDP), during peak ZIKV infection prevents synapse elimination and loss of adult hippocampal neurogenesis, without impacting CNS virologic control. Transcriptomic analyses of forebrain immune cells in MDP-treated mice revealed functional modulation of infiltrated monocytes and T cells, reducing their expression of pro-inflammatory cytokines, with limited effects on microglia, compared to controls. Notably, NOD2 activation in peripheral immune cells alone balances innate immune signals, preserving synapses, and increasing macrophage phagocytic capacities that do not target synapses. Our findings identify infiltrating Ly6CHi monocytes as key drivers of long-term cognitive dysfunction following ZIKV encephalitis and as potential therapeutic targets for limiting synapse loss. HighlightsO_LINOD2 activation via MDP phenotypically shifts monocyte subsets from inflammatory to anti-inflammatory during the acute phase of ZIKV encephalitis. C_LIO_LIShort-term MDP administration increases phagocytic machinery and reduces inflammatory cytokine/chemokine production within macrophage populations. C_LIO_LISynapse elimination is attenuated within the hippocampus of ZIKV-infected animals treated with MDP. C_LIO_LIMDP derived effects are mediated through peripherally derived immune cells. C_LI

Antibiotic exposure is a significant risk factor for Crohns disease, yet the tissue-specific consequences of antibiotic-driven dysbiosis remain poorly defined. Adherent-invasive Escherichia coli (AIEC), a pathobiont enriched in Crohns disease, expands following antibiotic treatment, but whether discrete mucosal niches support this expansion is unknown. Peyers patches are specialized lymphoid structures that coordinate mucosal immunity and are frequently associated with early disease lesions, suggesting they may represent a vulnerable site for pathobiont colonization. Here, we show that vancomycin disrupts the Peyers patch-associated microbiome, creating a permissive niche that is selectively exploited by AIEC and associated with focal inflammation. Antibiotic treatment markedly increased AIEC burden within Peyers patches. AIEC localized within the lymphoid follicle was accompanied by focal tissue pathology and a distinct cytokine signature. In contrast, expansion of resident E. coli in the absence of AIEC did not elicit comparable inflammation, indicating that the pathogenic traits of AIEC are required to trigger disease-relevant responses in this niche. Supporting this, genetic disruption of flagellin, long polar fimbriae, or antimicrobial peptide resistance in AIEC attenuated Peyers patch colonization or inflammation, revealing separable mechanisms governing niche access and immunopathology. Together, these findings identify Peyers patches as a previously unrecognized reservoir for antibiotic-driven AIEC expansion and define a localized host-microbe interaction that links dysbiosis to focal intestinal inflammation. These results provide a mechanistic framework for understanding how antibiotic exposure may precipitate site-specific pathology in Crohns disease. Further, these findings highlight that mucosal lymphoid tissues should be considered when evaluating microbiome-targeted therapeutic interventions in Crohns disease.

Epstein-Barr virus (EBV) infection is nearly ubiquitous in humans and has been associated with multiple sclerosis (MS) and other immune-mediated diseases, yet mechanisms by which EBV-infected B-cells influence distal tissues remain incompletely understood. Extracellular vesicles (EVs) mediate intercellular communication during viral infection, but their integrated viral and host cargo has not been comprehensively defined in EBV-transformed B-cells generated from naturally infected individuals. Here, we performed an unbiased multiomic characterization of small EVs (sEVs) released from spontaneous lymphoblastoid cell lines (SLCLs) derived from normal donors and patients with stable or active MS, transformed ex vivo by endogenous wild-type EBV. Using surface profiling, quantitative proteomics, whole-genome sequencing of EV-associated DNA, total stranded RNA sequencing, droplet digital PCR, and super-resolution microscopy, we mapped the protein, DNA, and RNA cargo of these vesicles and determined their host and viral origins. SLCL sEVs contained canonical tetraspanins, B-cell markers, and were free of detectable virions. Proteomics identified over 6,000 shared proteins and revealed enrichment of nucleic acid-binding and chromatin-associated proteins. Whole-genome sequencing demonstrated abundant EV-associated DNA comprising two distinct compartments: high-molecular weight, DNase-sensitive DNA associated with the vesicle corona and DNase-resistant, nucleosome-sized ([~]130-150 bp) DNA protected within vesicles. In both compartments, DNA was overwhelmingly host-derived and broadly distributed across the genome, whereas EBV genomic DNA was minimal. RNA sequencing identified diverse EBV transcripts, with striking enrichment of the viral long noncoding RNA EBER1 across all lines. Super-resolution imaging and ddPCR confirmed EBER1 incorporation within individual vesicles. Notably, EBER1 has been detected in MS brain tissue in prior studies, and our findings provide a plausible vesicle-mediated mechanism for dissemination of this viral lncRNA from EBV-infected B-cells to distal sites. These findings establish a foundational multiomic profile of sEVs from wild-type EBV-transformed B-cells and reveal export of host DNA and EBER1, with broad implications for viral immunobiology and intercellular signaling in MS and beyond. SIGNIFICANCE STATEMENTEpstein-Barr virus (EBV) infects over 90% of adults worldwide and is strongly linked to multiple sclerosis (MS). How EBV-infected B-cells may communicate with distant tissues, including the central nervous system (CNS), remains unclear. We provide the first integrated multiomic profile of extracellular vesicles released from B-cells transformed by endogenous, wild-type EBV. These vesicles are enriched in nucleosome-associated host DNA and the viral long noncoding RNA EBER1 but contain minimal viral DNA. Because EBER1 has been detected in MS brain tissue, our findings suggest a vesicle-mediated mechanism by which EBV-infected B-cells could deliver viral RNA to the CNS independently of infectious virus. These results establish a framework for understanding how EBV latency reshapes intercellular communication in immune-mediated disease.

The gastrointestinal tract (GI) is the largest environmental mucosal interface and is exposed to diverse commensal and pathogenic microbes. B cells are a prevalent immune component of the GI tract and its associated secondary lymphoid organs, yet we know little about the diversity and stability of distinct transcriptional programs that modulate B cell responses against different classes of pathogens or environmental perturbations. A subset of B cells defined by expression of the transcription factor T-bet, has been canonically associated with antiviral immunity through IgG production. However, the role of T-bet expressing B cells in mucosal tissues, where IgA responses predominate, is poorly understood. Here, we identify a population of intestinal T-bet+ B cells that, in the absence of overt perturbation, constitutes a minor fraction of intestinal associated B cells and undergoes continuous turnover. In contrast, during enteric viral infection with murine norovirus (MNV), T-bet B cells undergo a marked expansion, with T-bet expression stably maintained in the majority of virus-specific B cells, including IgG2c and IgA switched B cells. Moreover, virus-reactive IgG2c and IgA B cells arise independently rather than through sequential switching, and B cell intrinsic T-bet expression is required for effective germinal center responses but dispensable for IgA class switching. Moreover, T-bet expressing B cells are required for the generation of all MNV-specific circulating IgG and mucosal IgA, and for protection upon re-encounter with the virus. Together, these findings establish T-bet expressing B cells as a specialized B cell subset essential for mucosal immunity and protection against norovirus infection.

Therapeutic blockade of IL-17 and TNF can effectively treat inflammatory skin diseases such as hidradenitis suppurativa and psoriasis, yet the relative importance of the different cell types that respond to IL-17 and TNF remains unresolved. Keratinocytes are viewed as the dominant effector cells, whereas fibroblasts have recently emerged as important contributors. In mice, topical imiquimod induces IL-17- and TNF-dependent skin inflammation and is frequently used to model psoriasis. Here, we demonstrate that intradermal injection of recombinant IL-17 and TNF elicits skin inflammation with features of hidradenitis suppurativa, including a gene expression program that is distinct from psoriasis and imiquimod-induced inflammation. Single-cell transcriptomic network analysis identified dermal fibroblasts as the dominant cell communication hub in hidradenitis suppurativa and in mice injected with IL-17 and TNF. In contrast, fibroblasts and keratinocytes both show strong network involvement in psoriasis and in mice challenged with imiquimod. Cell-type-specific deletion of IL-17 receptor A in mice revealed that imiquimod-induced inflammation depends equally on IL-17 signaling in fibroblasts and keratinocytes, whereas inflammation induced by intradermal IL-17 and TNF only requires fibroblasts to recognize IL-17 and is independent of keratinocyte IL-17 sensing. Single-cell transcriptomic analysis of these conditional knockout mice further demonstrated that keratinocytes and fibroblasts activate divergent and disease-dependent transcriptional programs following activation by IL-17. Together, these findings introduce a new conceptual framework wherein IL-17 signaling is routed through distinct cellular and molecular pathways depending on disease context and establish complementary experimental systems for interrogating type 17 skin inflammation.

Fibroblasts are key organizers of tissue architecture and immune cell homeostasis, yet how they shape adaptive immune function within non-lymphoid tissues remains incompletely understood. CD8+ tissue-resident memory T cells (TRM) provide localized protection against pathogens and contribute to tumor control, but the microenvironmental signals that maintain their persistence and survival are poorly defined. Here, we identify fibroblast-derived TGF-{beta}3 as a conserved stromal niche factor that specifically sustains CD8+ TRM in both steady-state and disease settings. Across human single-cell cross-tissue atlases, CD8+ TRM preferentially correlated with fibroblast abundance in healthy barrier tissues and multiple tumor types, and TGFB3 emerged as a key fibroblast-enriched candidate mediator. In human and murine co-culture systems, fibroblast-derived TGF-{beta}3 promoted CD8 TRM-like differentiation in vitro. Using a novel genetic in vivo model, inducible fibroblast-specific deletion of Tgfb3 reduced CD8 TRM across barrier tissues at steady state and impaired antigen-specific CD8 TRM formation following viral infection. In tumor models, genetic loss or antibody mediated neutralization of TGF-{beta}3 impaired CD8 T cell residency and cytotoxicity, induced dysfunction via proteotoxic stress and apoptotic programs, and accelerated tumor growth. These findings provide mechanistic insight into the limited efficacy of pan-TGF-{beta} blockade in cancer therapy. Collectively, we describe a novel fibroblast-CD8 T cell axis mediated by TGF-{beta}3 that sustains residency and restrains proteotoxic stress in barrier tissues and tumors.

Chimeric antigen receptor (CAR)-T cell therapies can induce drug-free remission in systemic lupus erythematosus (SLE), but indiscriminate B cell targeting causes immunosuppression, unnecessary infections, and cytokine toxicities that preclude widespread use. Here, we overcome this by targeting the 9G4 idiotope, a shared structural feature of pathogenic B cell receptors encoded by the IGHV4-34 gene. We engineered anti-9G4 CAR-T cells and chimeric TCR-T cells to selectively eliminate autoreactive B cells while preserving protective immunity. Both platforms eradicated autoreactive B cells and autoantibodies in vitro and in vivo, spared normal B cells, and markedly reduced cytokine release compared to conventional CAR-T cells. This precision extended to cold agglutinin disease and lymphoma. These findings establish a framework for IGHV idiotope-directed cellular therapies for treating autoimmune and neoplastic diseases while preserving immune competence.