bluuebunny/crossref_metadata_2025_split huggingface dataset:
Interferon Gamma Induction of TH1-like Tregs Controls Anti-viral Responses
Authors: Gocher Angela Marie , Cui Jian , Szymczak-Workman Andrea , Vignali Kate , Latini Julianna N , Pieklo Gwen P , Avery Lyndsay , Cipolla Ellyse L , Huckestein Brydie R , Hedden Lee , Meisel Marlies , Alcorn John F , Kane Lawrence P , Workman Creg J , Vignali Dario A A 
Published: January 2023

Abstract
Regulatory T cells (Tregs) are an immunosuppressive cell population that inhibits immune cell function to maintain peripheral tolerance and homeostasis. We have previously shown that Treg-- response to interferon gamma (IFNγ) is required for response to cancer immunotherapy. We aimed to broaden these findings and determine the impact of Treg response to IFNγ and the IFNγ-inducing cytokine IL12, during viral infection. Mice with Treg-restricted deletion of the IFNγ receptor (IFNGR1, Ifngr1L/L Foxp3Cre-YFP) or the IL12 receptor β2 subunit (IL12Rβ2, Il12b2L/L Foxp3Cre-YFP), were infected with the acute (Armstrong), or chronic (Clone 13), strain of lymphocytic choriomeningitis virus (LCMV). Surprisingly, Treg response to IFNγ but not IL12 induced helper T cell 1 (TH1)-like polarization (expression of T-bet, CXCR3 and IFNγ) of Tregs in mice with chronic LCMV infection. Importantly, this effector-like TH1 Treg state was required for adequate suppression of effector T cells during LCMV infection. Additionally, during LCMV infection, Treg-restricted deletion of IFNGR1 not only prevented TH1-like polarization but also reverted these Tregs to a TH2-like state (expression of GATA-3, CCR4 and IL-4). scRNAseq of viral antigen-specific CD8+ T cells from Clone 13 infected mice showed that Ifngr1-deficient Tregs favored CD8+ T cell memory. These findings were confirmed in an Armstrong/Listeria monocytogenes-GP33 memory model. Further, Ifngr1 deletion from Tregs enhanced response to CD8+ T cell-mediated vaccination when challenged with Clone 13. These findings provide fundamental insight on how Tregs sense inflammatory cues from the environment (IFNγ) during viral infection to prevent overt tissue damage and shape the memory response.
Supported by grants from NIH (F32 CA247004-01, T32 CA082084, P01 AI108545, R35 CA263850, R01 CA203689, R01 DK130897, P30 DK120531, P30 CA047904, R01 HL107380, R01 CA206517, R01 AI138504)
TNF-α orchestrates the specific loss of tolerogenic dendritic cells during an acute maturation process in vivo
Authors: Bourque Jessica , Iberg Courtney , Fallahee Ian , Hawiger Daniel 
Published: January 2023

Abstract
The recognition of pathogen-associated molecular patterns (PAMPs) through the corresponding pattern recognition receptors (PRRs) induces activation of individual conventional dendritic cells (cDC) in a process often referred to as maturation that results in a change from tolerogenic to immunogenic functions. In the absence of pro-inflammatory conditions (steady state), tolerogenic cDC that are present in systemic peripheral lymphoid organs promote tolerance that limits subsequent immune responses. However, the fate of such tolerogenic cDC in the presence of specific pro-inflammatory signals remains unclear. Here, we report that instead of reprogramming individual tolerogenic cDC for new immunogenic functions, canonical inducers of maturation such as LPS initiated a burst of TNF-α production in these cDC, which resulted in their acute and specific death mediated by TNFR1. This ablation of individual tolerogenic cDC was enhanced by TNF-α produced by neighboring cDC, amplifying the initial innate stimulus. Overall, our results expand the understanding of the functions of TNF-α and TNFR1 during the maturation process and uncover that immunogenic maturation involves a dynamic remodeling of the cDC functional landscape by selectively ablating the tolerogenic cDC population to prevent an aberrant induction of tolerance. Therefore, our results may also have implications for devising new immunotherapeutic approaches.
This work was supported in part by grants from National Institute of Allergy and Infectious Diseases of the National Institutes of Health (R01AI113903) and National Multiple Sclerosis Society (RG-1902-33632), both to DH.
Tregs suppress antigen-specific CD8+ T cells in vivo by depleting pMHC-I complexes from Dendritic Cells
Authors: Mansoori Mohammad Nizam , Kamenyeva Olena , Kabat Juraj , Shevach Ethan M 
Published: January 2023

Abstract
T Regulatory cells (Tregs) regulate antigen-specific immune responses by using a variety of mechanisms. Recent studies have shown that antigen-specific Treg can suppress CD4+ effector T cells in vitro and in vivo by depleting peptide-MHC-II complexes from the DC surface. It remains unclear as to how Treg suppress CD8+ T effectors, particularly in vivo. To explore Treg-mediated suppression of CD8+ T cells, we generated CD4+ iTregs from OT-II mice specific for Ova323–339 in association with I-Ab and determined their capacity to suppress CD8+ T cells from OT-I mice specific for Ova257–264 (SIINFEKL) in association with H-2Kb. CD4+ OT-II iTreg suppressed the in vitro proliferation of OT-I T cells in the presence of DCs pulsed with both Ova323–339 and Ova257–264 or with DCs singly pulsed with each peptide. Interestingly, the expansion of OT-I CD8+ T cells in vivo was not suppressed by OT-II iTreg when the two peptides were presented on separately pulsed DCs, but was suppressed when both peptides were presented on the same DCs. OT-II Tregs depleted the Kb-SIINFEKL complexes from the DC surface in vitro by a process of trogocytosis only in the presence of their cognate antigen. The uptake of Kb-SIINFEKL complexes by OT-II Treg was not secondary to leakage of free SIINFEKL from the DC surface as OT-II Treg from MHC-I deficient mice were as efficient in uptake of pMHC-I as Treg from wild type mice. Multiphoton imaging microscopy in vivo reveals a close interaction between Tregs and CD8+ T cells suggesting that suppression of CD8+ T cells in vivo requires close proximity between Treg and responder CD8+ T cell may involve an artificial synapse between Treg and CD8+ T cell.
The activation of self-antigen specific T cell repertoire during acute tissue injury.
Authors: Shin Daniel Seong-Joo , Moon James J 
Published: January 2023

Abstract
Self-antigen specific T cells are prevalent in the mature adaptive immune repertoire. At steady state, these self-antigen specific T cells appear to be regulated through a combination of central and peripheral mechanisms of tolerance. Understanding how these self-antigen specific T cells escape tolerance to mediate autoimmune disease remains of significant scientific and clinical interest. Importantly, the mechanisms that determine whether the self-reactivity will exacerbate or regulate the ongoing immune response are unknown. Utilizing an experimental animal model where mice express a complex of three model T cell epitopes (2W, gp66, and OVA) in a tissue restricted manner, we identified and tracked the activation of endogenous polyclonal T cells that recognize these transgenic epitopes as self-antigens. Interestingly, in the context of acute injury to lung tissue of mice expressing the epitopes in the alveolar epithelium, we detect an asymmetric expansion of the self-antigen specific regulatory T cell (Treg) compartment while the conventional T cell populations remained static. These findings suggest that the self-antigen specific T cell repertoire is uniquely programmed to generate a protective response to acute tissue injury.
Disruption of IFNγ, PRF1, GZMB, or LYST results in reduced suppressive function in human CD8+ T cells
Authors: Vemulawada Chakrapani , Renavikar Pranav S , Crawford Michael P , Steward-Tharp Scott , Guo Hui , Jian Zhang , Karandikar Nitin J 
Published: January 2023

Abstract
Human CD8+ T-cells play an important immune regulatory role during autoimmune diseases, characterized by suppressing the activation and proliferation of pathogenic effector cells, mediated in part through cytotoxicity (PRF/GZM), IFNγ secretion or by Fas/FasL interactions. Recent studies from our group demonstrated that IL-12 exposure during in vitro differentiation of naïve CD8+ T-cells induced an immune-suppressive deficit, with Tc1 cells showing significant loss of suppressive ability against CD4+ T-cells. RNAseq analysis revealed that various cytokines, transcription factors, and other regulatory molecules were differentially regulated in these cells. In this study, we further investigated the importance of IFNγ-, PRF1-, GZMB- and LYST-associated pathways in the suppressive ability of CD8+ T-cells. Using the CRISPR-Cas9 RNP optimized transfection system in primary bulk human CD8+ T-cells. Knockout was confirmed through quantitative real-time PCR assays in all cases, combined with flow cytometry when possible, as well confirmation of indels at genomic target sites. We observed that knockout of each of the four molecules, but not the knockout of IL-4 or IL-5, resulted in significantly diminished suppressive ability, as measured through in vitro immune suppression assays. Interestingly, in some of the knockouts, we observed differential expression of non-target molecules indicating potential functional links between these pathways. Collectively, these results reveal a pivotal role for these pathways in CD8+ T cell-mediated immune suppression and provide important insights into the biology of human CD8+ T-cell-mediated suppression that could be targeted for immunotherapeutic intervention.
This work was supported, in part, by grant awards to NJK from the NIH (R01 AI121567) and VA.
Multiomic dynamic single-cell profiling of CAR T cell populations associated with efficacy
Authors: Rezvan Ali , Romain Gabrielle , Fathi Mohsen , Heeke Darren , Martinez-Paniagua Melisa , An Xingyue , Bandey Irfan N , Liadi Ivan , Cooper Laurence J N , Singh Harjeet , Bot Adrian , Neelapu Sattva , Varadarajan Navin 
Published: January 2023

Abstract
T-cell therapy with specificity redirected through chimeric antigen receptors (CARs) has shown efficacy for the treatment of hematologic malignancies. Although treatment with CAR T cell can result in high response rates, the properties of the cells that comprise the cellular infusion product, associated with clinical benefit are incompletely understood. We utilized a suite of high-throughput single-cell assays including single-cell RNA-sequencing (scRNA-seq); confocal microscopy; and Timelapse Imaging Microscopy In Nanowell Grids (TIMING). TIMING profiling of a cohort of 16 patients showed that persistent motility of T cells in the presence of tumor cells was associated with both serial killing capacity and polyfunctionality. Confocal microscopy on these same T cells revealed that persistent motility is linearly correlated with both mitochondrial volume and lysosomal number. ScRNA-seq demonstrated that T cells from responders were enriched in pathways related to T-cell proliferative capacity; interferon responses; and a distinct cluster of pathways related to actin cytoskeleton and migration. We employed a marker-free sorting strategy for enriching T cells with persistent motility. RNA-seq on sorted motile T cells showed an enrichment of the core motility signature. These motile T cells also demonstrated superior in vivo anti-leukemia efficacy in comparison to unsorted T cells. Lastly, we also confirmed the association with increased persistent motility and killing of CAR T cells across diverse CARs. In aggregate, our data identified that independent of CAR design or biomanufacturing, persistent motility serves as a selectable cell-intrinsic biomarker, desired in the bioactivity of expanded CAR+ T cells.
Amino acid F223 in neuroimmune semaphorin 4A is essential for stabilization of human regulatory T cells
Authors: Chapoval Svetlana P , Lee Mariah , Lemmer Aaron , Ajayi Oluwaseyi , Qi Xiulan , Neuwald Andrew F , Keegan Achsah Dorsey 
Published: January 2023

Abstract
Class IV semaphorin family members, Sema4A and Sema4D, displayed opposite effects on human Treg cells in vitro. In PBMC cultures, Sema4A potentiated the CD4+CD25+Foxp3+ cell numbers and increased CD25/Foxp3 expression while Sema4D inhibited them. Sema4A downregulated lung inflammation in allergen-induced experimental asthma whereas Sema4D was necessary for optimal allergic response. Allergen treatment decreased lung local and lymphoid tissue Treg cell numbers in Sema4A−/− mice and increased them in Sema4D−/− mice as compared to similarly treated WT counterparts. Sema4A and Sema4D competitively bind to Plexin B1 in microplate-based binding studies. Using Bayesian Partitioning with Pattern Selection (BPPS) and Structurally Interacting Pattern Residues’ Inferred Significance (SIPRIS), we identified Sema4A residues likely responsible for its functional specificity. We mutated the Sema4A residues M198 and F223 to alanine; notably, F223 in Sema4A corresponds to alanine in Sema4D. We evaluated their stimulatory activity toward human Treg cells and assayed them for binding to Plexin B1. The F223A mutant was unable to stimulate Treg stability in the in vitro PBMC cultures despite binding Plexin B1 with an affinity similar to WT and M198A proteins. This research is a first step in generating potent mutant Sema4A molecules with stimulatory function for Treg cells with a view to designing novel immunotherapeutics for asthma.
This work was supported by NIH grant R01AI 122631 and the Merit Review Award Number I01 BX001850 from the US Department of Veterans Affairs Biomedical R&D Service (both awarded to A.D.K.) and by NIH-NIGMS grant R01GM125878 to A.F.N.
Mettl14-dependent m6A modification controls iNKT cells development and function
Authors: Cao Liang , Morgun Eva , Genardi Samantha , Visvabharathy Lavanya , Huang Haochu , Wang Chyung-Ru 
Published: January 2023

Abstract
N6-Methyladenosine (m6A), the most common form of RNA modification, controls CD4+ T cell homeostasis by targeting the IL-7/STAT5/SOCS signaling pathways. The role of m6A modification in unconventional T cell development remains unknown. Using mice with T cell-specific deletion of RNA methyltransferase METTL14 (T-Mettl14−/−), we demonstrated that m6A modification was indispensable for iNKT cell homeostasis. Loss of METTL14-dependent m6A modification led to the upregulation of p53-mediated apoptosis in double positive thymocytes, which in turn decreased distal Va-Ja gene rearrangements, including the invariant Va14-Ja18 TCR, resulting in a drastic reduction of iNKT numbers in the thymus and periphery. Residual T-Mettl14−/− iNKT cells exhibited increased apoptosis, impaired maturation, and decreased responsiveness to IL-2/IL-15 and TCR stimulation. Furthermore, METTL14 knockdown in mature iNKT cells diminished their cytokine production, correlating with increased Cish expression and decreased TCR signaling. Collectively, our study highlights a critical role for METTL14-dependent-m6A modification in iNKT cell development and function.
This work was supported by NIH grants R01 AI41083 and R01 AI057460 to C.-R.W.
The Role of HnRNPA1 in T Cell-Mediated Gut Tolerance
Authors: White Tristan L , Gable Matthew , Jin Ye , Morel Penelope A 
Published: January 2023

Abstract
Our previous research showed that Akt phosphorylation of the RNA-binding protein, (RBP) heterogeneous nuclear ribonucleoprotein (hnRNP) A1, is dependent on TCR signal strength, and occurs under conditions of induced T regulatory (Treg) cell differentiation. We have shown hnRNPA1 is required for optimal Treg differentiation by performing knockdown experiments, and our present research is focused on identifying a role for Akt phosphorylation in hnRNPA1 function. HnRNPA1 is phosphorylated by Akt at S199 and our lab has generated a new mutant mouse model, hnRNPA1-S199A (mA1), in which hnRNPA1 will no longer be Akt-phosphhorylated. Our initial characterization of mA1 mouse model failed to reveal major abnormalities in immune cells populations at steady state except for changes in the Treg proportion in the mesenteric lymph nodes. To investigate the effect of the mA1 on the development of oral tolerance we used an adoptive transfer model in which OT-II mA1 or OT-II WT naïve T cells were transferred into congenic strains and then the mice were given OVA food. Results indicate that mA1 OT-II T cells have a proliferation defect in vivo and we also observed a significant reduction in the induction of OVA-specific Treg in the mA1 T cells. Similar results were obtained when adoptively transferred OT-II mA1 T cells were challenged with limiting doses of specific peptide, given iv. RNASeq analysis of the transferred mA1 and WT T cells is being performed. These results suggest that Akt phosphorylation of hnRNPA1 is necessary for T cell response to low dose antigen and Treg induction in vivo. Our studies identify a new mechanism by which RBP influence T cell proliferation and Treg differentiation which has implications for tolerance to food antigens in the gut.
Supported by NIH (F31-AI152320-02) NIH (R01-AI125513-03)
Role of transcription factor RUNX1 in the maintenance of T helper cell epigenome and rapid recall response
Authors: Korinfskaya Svetlana , Kotliar Michael , Katko Alexander , Hildeman David , Miraldi Emily R , Barski Artem 
Published: January 2023

Abstract
Memory T cells exhibit an accelerated response after a second encounter with the same antigen. In our previous studies, we showed that the ability to produce effector molecules within minutes of the secondary exposure to antigen (“rapid recall response”) is associated with the epigenetic state of the rapid recall genes. This memory epigenome is established during the primary T cell encounter with its cognate antigen. To maintain a “rapid recall response” even when antigen is eliminated, resting memory T cells would have to maintain open chromatin signature for years despite episodic homeostatic T cell proliferation. However, the mechanisms underlying the long-term maintenance of this epigenetic pattern remains unclear. Here, we used 10X Multiome (scRNA-seq and scATAC-seq obtained from the same cells) analysis to show that the association between the chromatin state of memory cells and their cytokine expression upon activation is Th lineage specific.
Bioinformatic analysis showed that open chromatin specific for memory but not naïve CD4 T cells is enriched with motifs of constitutively expressed transcription factors RUNX, SP1, and ETS. Our preliminary Cut&Tag data in total human resting CD4 T cells demonstrates that RUNX1 binds DNA close to T helper cytokines genes (IL3, IL4, IL9, IL10, IL13, IFNG) at “memory-specific” ATAC-seq regions, suggesting the role of RUNX1 in maintaining this “rapid recall” epigenome. Together with the dynamic model of transcriptome and epigenome rearrangements this data could help us to understand how memory T cells establish a platform that enables and maintains their unique ability to respond more rapidly to subsequent encounters with antigen.
Supported by R01AI153442-01A1 from NIAID, NIH
Epigenome accessibility changes before and after activation reveal distinct and progressive differentiation for human memory T cell subsets
Authors: Rose James R , Rahmberg Andrew R , Powell Michael D , Scharer Christopher D , Boss Jeremy M 
Published: January 2023

Abstract
Memory T cells (MTC), an indispensable part of adaptive immune memory, have historically been categorized by the cell surface proteins CCR7 and CD45RA into specialized subtypes (TCM, TEM, and TEMRA), each with unique functions. However, the epigenetic characteristics that distinguish the MTC subsets as well as the gene regulatory networks governing each MTC subsets’ response to activation remain poorly understood. Here we define and categorize the transcriptional and epigenetic differences of MTC and their respective primary subsets found in human blood, both in a resting state and after ex-vivo stimulation. Resting TCM were found to be relatively more similar to naïve cells in both CD4 and CD8 lineages, while TEM exhibited greater numbers of differentially expressed genes (DEG) and alterations to chromatin accessibility. Differentially accessible regions (DAR) discerning memory subsets contained binding motifs for factors thought to regulate memory formation from the bZIP, T-box and HMG families, as well as sites for novel bHLH factors MSC and AHR that may function in MTC development. Examining the effect of stimulation on MTC gene expression and chromatin identified unique DAR and DEG modules, some of which may have initially been primed by previous activation events in naive progenitors. Primed DAR were correlated with augmented expression of important genes in MTC after stimulation, suggesting an epigenetic mechanism of regulation. Ultimately these results describe the accumulation of epigenetic alterations during primary activation and during memory development that distinguish MTC from naïve T cells, enable differentiation into distinct subsets, and influence how MTC respond to secondary activation.
Supported by grants from NIH (RO1 AI113021,T32 GM0008490)
Human T cells in barrier sites exhibit site-specific characteristics and clonal compartmentalization
Authors: Caron Daniel P , Poon Maya M L , Wang Zicheng , Meng Wenzhao , Lam Nora , Szabo Peter A , Wells Steven B , Thapa Puspa , Dogra Pranay , Lee Brian , Kubota Masaru , Matsumoto Rei , Rahman Adeeb , Luning Prak Eline T , Sims Peter , Shen Yufeng , Farber Donna L 
Published: January 2023

Abstract
The skin, gut, and lung are critical physical and immune barriers that protect from pathogen entry. Within these sites, T cell memory is largely maintained by populations of tissue resident memory T cells (TRMs), capable of rapid protective responses. The degree of interconnection between these TRMs from various sites of residence is poorly understood, particularly in humans. Here, we analyze how tissue specificity affects transcriptional heterogeneity and the T cell receptor (TCR) repertoire across 9 tissue sites within individual donors, including barrier sites (skin, gut, and lung) and associated lymph nodes, as well as blood and lymphoid organs (blood, bone marrow, spleen). Cytometry by time-of-flight (CyTOF) and single cell RNA sequencing reveal phenotypic and transcriptomic features unique to skin or gut T cells, which are distinct from T cells in lymphoid sites or blood. These results were reflected in both bulk sequencing of TRBV gene rearrangement and single cell TCR sequencing, where skin and gut T cells exhibited site-specific clonal expansions that were not present in other sites. Conversely, lung T cells showed clonal overlap and transcriptional similarity to T cells in lymphoid and blood rich sites. Site-specific expansions were mediated by TRMs, while clones disseminated across tissues exhibit large clonal expansions and a CD8 terminal effector (TEMRA) phenotype. Together, these results reveal that TRMs in barrier sites are maintained in situ, while memory T cells in the lung, blood-rich and lymphoid sites are more interconnected. Blood T cells are not representative of barrier T cell clonal space. These results have important implications for monitoring and promoting barrier immunity through site-specific targeting.
Supported by grants from NIH (P01 AI106697)