bluuebunny/crossref_metadata_2025_split huggingface dataset:
Constitutively Active Aryl Hydrocarbon Receptor Expressed Specifically in T-Lineage Cells Causes Thymus Involution and Suppresses the Immunization-Induced Increase in Splenocytes
Authors: Nohara Keiko , Pan Xiaoqing , Tsukumo Shin-ichi , Hida Azumi , Ito Tomohiro , Nagai Haruko , Inouye Kaoru , Motohashi Hozumi , Yamamoto Masayuki , Fujii-Kuriyama Yoshiaki , Tohyama Chiharu 
Published: April 2014

AbstractThe aryl hydrocarbon receptor (AhR) is a transcription factor belonging to the basic helix-loop-helix-PER-ARNT-SIM superfamily. Xenobiotics, such as 2,3,7,8-tetrachlorodibenzo-p-dioxin, bind the receptor and trigger diverse biological reactions. Thymocyte development and T cell-dependent immune reactions are sensitive targets of AhR-dependent 2,3,7,8-tetrachlorodibenzo-p-dioxin toxicity. However, the exact role of the AhR in T cells in animals exposed to exogenous ligands has not been clarified because indirect effects of activated AhR in other cell types cannot be excluded. In this study, we generated transgenic (Tg) mice expressing a constitutively active mutant of AhR under the regulation of a T cell-specific CD2 promoter to examine AhR function in T cells. The mRNAs of the constitutively active mutant of AhR and an AhR-induced gene, CYP1A1, were expressed in the thymus and spleen of the Tg mice. The transgene expression was clearly detected in the thymocytes, CD4, and CD8 T cells, but not in the B cells or thymus stromal cells. These Tg mice had a decreased number of thymocytes and an increased percentage of CD8 single-positive thymocytes, but their splenocytes were much less affected. By contrast, the increase in number of T cells and B cells taking place in the spleen after immunization was significantly suppressed in the Tg mice. These results clearly show that AhR activation in the T-lineage cells is directly involved in thymocyte loss and skewed differentiation. They also indicate that AhR activation in T cells and not in B cells suppresses the immunization-induced increase in both T cells and B cells.
Overlapping Signaling Pathways of Sphingosine 1-Phosphate and TGF-β in the Murine Langerhans Cell Line XS52
Authors: Radeke Heinfried H. , von Wenckstern Henrik , Stoidtner Kirsten , Sauer Bettina , Hammer Stefanie , Kleuser Burkhard 
Published: April 2014

AbstractTGF-β has been defined as a key mediator for the induction and maintenance of immunological tolerance. Concomitantly, it is essential for homeostasis of specialized epithelial dendritic cells, namely, Langerhans cells (LC). Our data reveal that TGF-β induces migration of the immature LC, XS52, a cell line expressing the signaling components, TGF-β type I and II receptors and Smad2, 3, and 4 mRNA. TGF-β stimulation induced transient Smad3/4 oligomerization and Smad3/DNA binding. Antisense oligonucleotides (ASO) targeting Smad3 abrogated TGF-β-induced XS52 chemotaxis, proving the involvement of this Smad protein in the TGF-β-dependent migration. In contrast, the typical CCR6-dependent chemotaxis of immature LC induced by CCL20/MIP-3α was not affected by Smad3 ASO. Most notably, we also identified the lysophospholipid sphingosine 1-phosphate (S1P) as a potent chemoattractant for immature LC, which expressed mRNA transcripts of lysophospholipid receptors S1P1–4. Additional experiments with specific ASO showed that the Gαi-coupled receptors S1P1 and S1P3 were dominantly involved in the S1P-induced migration. In contrast, lysophosphatidic acid (LPA), also binding to members of the lysophospholipid receptor family, failed to induce XS52 migration. Intriguingly, we raised evidence that TGF-β and S1P signal transduction pathways are indeed overlapping, as S1P augmented Smad activation and targeted DNA binding with kinetics comparable to TGF-β. Finally, S1P failed to stimulate XS52 chemotaxis when Smad3 protein expression was abrogated. Thus, our data indicate a cross-communication between S1P and TGF-β signaling that might be relevant for more than only migratory activities of immature LC.
Generation of Antigen-Specific, Foxp3-Expressing CD4+ Regulatory T Cells by Inhibition of APC Proteosome Function
Authors: Cong Yingzi , Konrad Astrid , Iqbal Nuzhat , Hatton Robin D. , Weaver Casey T. , Elson Charles O. 
Published: April 2014

Abstract
We tested the hypothesis that immature APC, whose NF-κB-signaling pathway and thus maturation was blocked by the proteosome inhibitor benzyloxycarbonyl-isoleucyl-glutamyl(O-tert-butyl)-alanyl-leucinal (PSI), could be a source of Ag-specific regulatory T (Treg) cells. DO11.10 CD4+ T cells that were incubated with Ag- and PSI-pulsed APC proliferated poorly, produced less IL-2, IFN-γ, and IL-10 in secondary cultures, and inhibited the response of both naive and memory CD4+ T cells stimulated by Ag-pulsed APC. The generation of PSI-APC Treg cells required IL-10 production by APC. PSI-APC Treg cell inhibition required cell-cell contact but not IL-10 or TGF-β. Addition of IL-2 did not reverse, but Ab to CTLA-4 did reverse partially the inhibitory effect. Depletion of CD25+ T cells before initial culture with PSI-APC did not affect Treg generation. PSI-APC Treg cells expressed high levels of Foxp3, inhibited proliferation of naive DO11.10 T cells in vivo, and abrogated colitis driven by a memory Th1 response to bacterial-associated Ag. We conclude that NF-κB-blocked, immature APC are able to induce the differentiation of Treg cells that can function in vitro and in vivo in an Ag-specific manner.
Synergistic Effect of IL-2, IL-12, and IL-18 on Thymocyte Apoptosis and Th1/Th2 Cytokine Expression
Authors: Rodriguez-Galán Maria Cecilia , Bream Jay H. , Farr Andrew , Young Howard A. 
Published: April 2014

AbstractIn the periphery, IL-18 synergistically induces the expression of the Th1 cytokine IFN-γ in the presence of IL-12 and the Th2 cytokines IL-5 and IL-13 in the presence of IL-2. Although the expression of these cytokines has been described in the thymus, their role in thymic development and function remains uncertain. We report here that freshly isolated thymocytes from C57BL/6 and BALB/c mice stimulated in vitro with IL-2-plus-IL-18 or IL-12-plus-IL-18 produce large amounts of IFN-γ and IL-13. Analysis of the thymic subsets, CD4−CD8− (DN), CD4+CD8+, CD4+CD8−, and CD4−CD8+ revealed that IL-18 in combination with IL-2 or IL-12 induces IFN-γ and IL-13 preferentially from DN cells. Moreover, DN2 and DN3 thymocytes contained more IFN-γ+ cells than cells in the later stage of maturation. Additionally, IL-18 in combination with IL-2 induces CCR4 (Th2-associated) and CCR5 (Th1-associated) gene expression. In contrast, IL-18-plus-IL-12 specifically induced CCR5 expression. The IL-2-plus-IL-18 or IL-12-plus-IL-18 effect on IFN-γ and IL-13 expression is dependent on Stat4 and NF-κB but independent of Stat6, T-bet, or NFAT. Furthermore, IL-12-plus-IL-18 induces significant thymocyte apoptosis when expressed in vivo or in vitro, and this effect is exacerbated in the absence of IFN-γ. IL-12-plus-IL-18-stimulated thymocytes can also induce IA-IE expression on cortical and medullary thymic epithelial cells in an IFN-γ-dependent manner. Thus, the combination of IL-2, IL-12, and IL-18 can induce phenotypic and functional changes in thymocytes that may alter migration, differentiation, and cell death of immature T cells inside the thymus and potentially affect the Th1/Th2 bias in peripheral immune compartments.
IgH V-Region Sequence Does Not Predict the Survival Fate of Human Germinal Center B Cells
Authors: Jackson Stephen M. , Capra J. Donald 
Published: April 2014

AbstractGerminal center (GC) B cell survival fate is governed in part by the outcome of successful/failed BCR-mediated interactions with accessory cells. However, the extent to which the BCR primary sequence influences such interactions is not fully understood. Over 1000 IgVH4 family cDNAs were sequenced from living (annexin V−) and apoptotic (annexin V+ or from within tingible body macrophages) GC B cell fractions from seven tonsils. Results surprisingly demonstrate that living and dying GC B cells do not significantly differ in IgVH, D, or JH gene segment use; HCDR3 length or positive charge; or mutation frequency. Additionally, equivalent IgH cDNA sequences were identified in both fractions, suggesting that BCR sequence alone is an unreliable predictor of GC B cell survival.
EBV-Induced Gene 3 Transcription Is Induced by TLR Signaling in Primary Dendritic Cells via NF-κB Activation
Authors: Wirtz Stefan , Becker Christoph , Fantini Massimo C. , Nieuwenhuis Edward E. , Tubbe Ingrid , Galle Peter R. , Schild Hans-Jörg , Birkenbach Mark , Blumberg Richard S. , Neurath Markus F. 
Published: April 2014

Abstract
The EBV-induced gene 3 (EBI3) is expressed in dendritic cells (DCs) and part of the cytokine IL-27 that controls Th cell development. However, its regulated expression in DCs is poorly understood. In the present study we demonstrate that EBI3 is expressed in splenic CD8−, CD8+, and plasmacytoid DC subsets and is induced upon TLR signaling. Cloning and functional analysis of the EBI3 promoter using in vivo footprinting and mutagenesis showed that stimulation via TLR2, TLR4, and TLR9 transactivated the promoter in primary DCs via NF-κB and Ets binding sites at −90 and −73 bp upstream of the transcriptional start site, respectively. Furthermore, we observed that NF-κB p50/p65 and PU.1 were sufficient to transactivate the EBI3 promoter in EBI3-deficient 293 cells. Finally, induced EBI3 gene expression in DCs was reduced or abrogated in TLR-2/TLR4, TLR9, and MyD88 knockout mice, whereas both basal and inducible EBI3 mRNA levels in DCs were strongly suppressed in NF-κB p50-deficient mice. In summary, these data suggest that EBI3 expression in DCs is transcriptionally regulated by TLR signaling via MyD88 and NF-κB. Thus, EBI3 gene transcription in DCs is induced rapidly by TLR signaling during innate immune responses preceding cytokine driven Th cell development.
Involvement of TNF and NF-κB in the Transcriptional Control of Cyclooxygenase-2 Expression by IFN-γ in Macrophages
Authors: Vila-del Sol Virginia , Fresno Manuel 
Published: April 2014

AbstractIFN-γ induces cyclooxygenase (COX)-2 expression and PG production in mouse macrophage cells. IFN-γ activates COX-2 promoter-driven transcription. Deletion of the IFN sequence regulatory element (ISRE) I −1541/−1522 and ISRE II −1215/−1206 sites of the mouse COX-2 promoter minimally decrease this IFN-γ induction. In contrast, deletion of the −965/−150 region from the COX-2 promoter abrogated IFN-γ induction. In this region a NF-κB site has been described and mutation of this site impairs the induction of the full COX-2 promoter by IFN-γ. Moreover, IFN-γ induction of the COX-2 promoter was also strongly reduced by transfection of plasmid encoding the NF-κB inhibitor, IκBα. Interestingly, IFN-γ induction of the COX-2 and PGE2 synthesis was absent in macrophages from TNF−/− mice, and neutralizing anti-TNF Abs inhibited COX-2 promoter induction by IFN-γ in RAW 264.7 macrophages. Moreover, NF-κB activity was induced late after stimulation with IFN-γ correlating with the effect of autocrine TNF, and this NF-κB activation was absent in macrophages from TNF−/− mice. Taken together our results suggest a model in which IFN-γ-induced TNF activates NF-κB, which is required for full COX-2 expression.
The Igκ3′ Enhancer Is Activated by Gradients of Chromatin Accessibility and Protein Association
Authors: McDevit Daniel C. , Perkins Leslie , Atchison Michael L. , Nikolajczyk Barbara S. 
Published: April 2014

AbstractThe Igκ locus is recombined following initiation of a signaling cascade during the early pre-B stage of B cell development. The Ig κ3′ enhancer plays an important role in normal B cell development by regulating κ locus activation. Quantitative analyses of κ3′ enhancer chromatin structure by restriction endonuclease accessibility and protein association by chromatin immunoprecipitation in a developmental series of primary murine B cells and murine B cell lines demonstrate that the enhancer is activated progressively through multiple steps as cells mature. Moderate κ3′ chromatin accessibility and low levels of protein association in pro-B cells are increased substantially as the cells progress from pro- to pre-B, then eventually mature B cell stages. Chromatin immunoprecipitation assays suggest transcriptional regulators of the κ3′ enhancer, specifically PU.1 and IFN regulatory factor-4, exploit enhanced accessibility by increasing association as cells mature. Characterization of histone acetylation patterns at the κ3′ enhancer and experimental inhibition of histone deacetylation suggest changes therein may determine changes in enzyme and transcription factor accessibility. This analysis demonstrates κ activation is a multistep process initiated in early B cell precursors before Igμ recombination and finalized only after the pre-B cell stage.
Proteolytic Regulation of Activated STAT6 by Calpains
Authors: Zamorano Jose , Rivas Maria Dolores , Setien Fernando , Perez-G Moises 
Published: April 2014

AbstractThe transcription factor STAT6 plays an important role in cell responses to IL-4. Its activation is tightly regulated. STAT6 phosphorylation is associated with JAKs, whereas dephosphorylation is associated with specific phosphatases. Several studies indicate that proteases can also regulate STAT6. The aim of this study was to investigate the nature of these proteases in mouse T cell lines. We found that STAT6 was degraded in cell extracts by calcium-dependent proteases. This degradation was specifically prevented by calpain inhibitors, suggesting that STAT6 was a target for these proteases. This was supported by the cleavage of STAT6 by recombinant calpains. The proteolytic regulation of STAT6 was more complex in vivo. Calcium signaling was not sufficient to induce STAT6 degradation. However, treatment of IL-4-stimulated cells with calcium ionophores resulted in the absence of phosphorylated STAT6. This effect correlated with the loss of STAT6 protein and was prevented by calpain inhibitors. Cytoplasmic calpains seemed to be responsible for STAT6 degradation. Calpains can target signaling proteins; in this study we found that they can negatively regulate activated STAT6.
Localization of Src Homology 2 Domain-Containing Phosphatase 1 (SHP-1) to Lipid Rafts in T Lymphocytes: Functional Implications and a Role for the SHP-1 Carboxyl Terminus
Authors: Fawcett Vicki C. J. , Lorenz Ulrike 
Published: April 2014

Abstract
The protein tyrosine phosphatase Src homology 2 domain-containing phosphatase 1 (SHP-1) has previously been shown to be a negative regulator of signaling mediated via the TCR. A growing body of evidence indicates that the regulated localization of proteins within certain membrane subdomains, referred to as lipid rafts, is important for the successful transduction of signaling events downstream of the TCR. However, considerably less is known about the localization of negative regulators during these lipid raft-dependent signaling events. In this study we have investigated the subcellular localization of SHP-1 and its role in regulation of TCR-mediated signaling. Our studies demonstrate that in a murine T cell hybridoma as well as in primary murine thymocytes, a fraction of SHP-1 localizes to the lipid rafts, both basally and after TCR stimulation. Interestingly, although SHP-1 localized in the nonraft fractions is tyrosine phosphorylated, the SHP-1 isolated from the lipid rafts lacks the TCR-induced tyrosine phosphorylation, suggesting physical and/or functional differences between these two subpopulations. We identify a requirement for the C-terminal residues of SHP-1 in optimal localization to the lipid rafts. Although expression of SHP-1 that localizes to lipid rafts potently inhibits TCR-mediated early signaling events and IL-2 production, the expression of lipid raft-excluded SHP-1 mutants fails to elicit any of the inhibitory effects. Taken together these studies reveal a key role for lipid raft localization of SHP-1 in mediating the inhibitory effects on T cell signaling events.
In Situ Diversification of the Antibody Repertoire in Chronic Lyme Arthritis Synovium
Authors: Ghosh Srimoyee , Steere Allen C. , Stollar B. David , Huber Brigitte T. 
Published: April 2014

AbstractLyme arthritis is initiated by the tick-borne spirochete, Borrelia burgdorferi. In a subset of patients, symptoms do not resolve in response to standard courses of antibiotics. Chronic joint inflammation may persist despite spirochetal killing, suggesting an autoimmune etiology. The pathogenic mechanisms that sustain chronic Lyme arthritis have not been fully elucidated, although T cells are believed to play a role. The synovial lesion contains elements of a peripheral lymph node, with lymphoid aggregates, plasma cells and follicular dendritic cells. An analysis of activated cells at the site of injury could yield clues regarding the nature of the response and the identity of potential autoantigens. Using laser-capture microdissection, we have isolated plasma cells from the joint tissue of chronic Lyme arthritis patients who underwent synovectomy. Expressed Ig V regions were amplified by RT-PCR. A majority of isolated cells expressed γ H chains, which is indicative of a class-switched response. There were a large number of nucleotide substitutions from germline, with a higher fraction of replacement mutations in the CDRs, suggesting a process of Ag-driven selection. We have recovered clonal clusters of cells containing identical junctions and V(D)J rearrangements. Sequence analysis reveals a hierarchy of shared somatic mutations between members of a given clone. Intraclonal diversity among plasma cells of close physical proximity points toward an ongoing process of diversification and affinity maturation, possibly driven by the chronic presence of an autoantigen.
The Two Major Oligomeric Forms of Human Mannan-Binding Lectin: Chemical Characterization, Carbohydrate-Binding Properties, and Interaction with MBL-Associated Serine Proteases
Authors: Teillet Florence , Dublet Bernard , Andrieu Jean-Pierre , Gaboriaud Christine , Arlaud Gérard J. , Thielens Nicole M. 
Published: April 2014

AbstractMannan-binding lectin (MBL) is an oligomeric C-type lectin assembled from homotrimeric structural units that binds to neutral carbohydrates on microbial surfaces. It forms individual complexes with MBL-associated serine proteases (MASP)-1, -2, -3 and a truncated form of MASP-2 (MAp19) and triggers the lectin pathway of complement through MASP-2 activation. To characterize the oligomerization state of the two major MBL forms present in human serum, both proteins were analyzed by mass spectrometry. Mass values of 228,098 ± 170 Da (MBL-I) and 304,899 ± 229 Da (MBL-II) were determined for the native proteins, whereas reduction of both species yielded a single chain with an average mass of 25,340 ± 18 Da. This demonstrates that MBL-I and -II contain 9 and 12 disulfide-linked chains, respectively, and therefore are trimers and tetramers of the structural unit. As shown by surface plasmon resonance spectroscopy, trimeric and tetrameric MBL bound to immobilized mannose-BSA and N-acetylglucosamine-BSA with comparable KD values (2.2 and 0.55 nM and 1.2 and 0.96 nM, respectively). However, tetrameric MBL exhibited significantly higher maximal binding capacity and lower dissociation rate constants for both carbohydrates. In contrast, no significant difference was detected for binding of the recombinant MASPs or MAp19 to immobilized trimeric or tetrameric MBL. As shown by gel filtration, both MBL species formed 1:2 complexes with MASP-3 or MAp19. These results provide the first precise analysis of the major human MBL oligomers. The oligomerization state of MBL has a direct effect on its carbohydrate-binding properties, but no influence on the interaction with the MASPs.
Interactions between NKG2x Immunoreceptors and HLA-E Ligands Display Overlapping Affinities and Thermodynamics
Authors: Kaiser Brett K. , Barahmand-pour Fariba , Paulsene Wendy , Medley Scott , Geraghty Daniel E. , Strong Roland K. 
Published: April 2014

AbstractThe NKG2x/CD94 family of C-type lectin-like immunoreceptors (x = A, B, C, E, and H) mediates surveillance of MHC class Ia cell surface expression, often dysregulated during infection or tumorigenesis, by recognizing the MHC class Ib protein HLA-E that specifically presents peptides derived from class Ia leader sequences. In this study, we determine the affinities and interaction thermodynamics between three NKG2x/CD94 receptors (NKG2A, NKG2C, and NKG2E) and complexes of HLA-E with four representative peptides. Inhibitory NKG2A/CD94 and activating NKG2E/CD94 receptors bind HLA-E with indistinguishable affinities, but with significantly higher affinities than the activating NKG2C/CD94 receptor. Despite minor sequence differences, the peptide presented by HLA-E significantly influenced the affinities; HLA-E allelic differences had no effect. These results reveal important constraints on the integration of opposing activating and inhibitory signals driving NK cell effector functions.
Hagfish Leukocytes Express a Paired Receptor Family with a Variable Domain Resembling Those of Antigen Receptors
Authors: Suzuki Takashi , Shin-I Tadasu , Fujiyama Asao , Kohara Yuji , Kasahara Masanori 
Published: April 2014

Abstract
Jawed vertebrates are equipped with TCR and BCR with the capacity to rearrange their V domains. By contrast, jawless vertebrates, represented by hagfish and lampreys, apparently lack such receptors. We describe in this study a family of hagfish genes carrying a single V-type domain resembling those of TCR/BCR. This multigene family, which we call agnathan paired receptors resembling Ag receptors (APAR), is expressed in leukocytes and predicted to encode a group of membrane glycoproteins with organizations characteristic of paired Ig-like receptors, consisting of activating and inhibitory forms. APAR has a J region in its V-type domain, and its V and J regions are encoded in a single exon. Thus, APAR is a member of the emerging families of diversified, innate immune-type receptors with TCR/BCR-like V-type domains and has many of the features expected for a primordial TCR/BCR-like receptor. The extracellular domain of APAR may be descended from a V-type domain postulated to have acquired recombination signal sequences in a jawed vertebrate lineage.
Intracellular Salmonella Inhibit Antigen Presentation by Dendritic Cells
Authors: Cheminay Cédric , Möhlenbrink Annette , Hensel Michael 
Published: April 2014

Abstract
Dendritic cells (DC) are important APCs linking innate and adaptive immunity. During analysis of the intracellular activities of Salmonella enterica in DC, we observed that viable bacteria suppress Ag-dependent T cell proliferation. This effect was dependent on the induction of inducible NO synthase by DC and on the function of virulence genes in Salmonella pathogenicity island 2 (SPI2). Intracellular activities of Salmonella did not affect the viability, Ag uptake, or maturation of DC, but resulted in reduced presentation of antigenic peptides by MHC class II molecules. Increased resistance to reinfection was observed after vaccination of mice with SPI2-deficient Salmonella compared with mice vaccinated with SPI2-proficient Salmonella, and this correlated with an increased amount of CD4+ as well as CD8+ T cells. Our study is the first example of interference of an intracellular bacterial pathogen with Ag presentation by DC. The subversion of DC functions is a novel strategy deployed by this pathogen to escape immune defense, colonize host organs, and persist in the infected host.