bluuebunny/crossref_metadata_2025_split huggingface dataset:
Fyn Membrane Localization Is Necessary to Induce the Constitutive Tyrosine Phosphorylation of Sam68 in the Nucleus of T Lymphocytes
Authors: Lang Valérie , Semichon Monique , Michel Frédérique , Brossard Cédric , Gary-Gouy Hélène , Bismuth Georges 
Published: December 2022

AbstractA close relationship between Sam68, a tyrosine and proline-rich RNA-binding protein, and Src protein tyrosine kinases (PTK) has already been established, also in T lymphocytes. A constitutive phosphorylation of the molecule has also been documented in various transformed T cells, which probably reflects an increased expression of PTK of the Src family. Using the hybridoma T cell line, T8.1, or Jurkat T cells, we investigated the respective contribution of the two Src kinases Fyn and Lck, expressed in T cells, in this phenomenon. By overexpressing the two proteins, we show that the constitutive phosphorylation of Sam68 in vivo directly correlates with cellular Fyn levels, but not with Lck expression, despite the capacity of the PTK to strongly phosphorylate the molecule in vitro. Overexpressed Fyn is mainly localized at the cell membrane. We find that Sam68 phosphorylation, including in the nuclear fraction in which the molecule is predominantly expressed, is lost with a delocalized Fyn mutant deleted of its N-terminal membrane-anchoring domain. Finally, we demonstrate, using a construct encoding a Sam68 molecule without its nuclear localization signal, that nuclear expression of Sam68 is not required for phosphorylation. We conclude that the constitutive phosphorylation of Sam68 in T cells is a Fyn-dependent process occurring in a cell-membrane compartment from which phospho-Sam68 molecules can thereafter accumulate into the nucleus.
A Central Role for αβ T Cells in the Pathogenesis of Murine Lupus
Authors: Seery John P. , Wang Eddie C. Y. , Cattell Victoria , Carroll Joseph M. , Owen Michael J. , Watt Fiona M. 
Published: December 2022

AbstractWe have previously shown that female transgenic mice expressing IFN-γ in the epidermis, under the control of the involucrin promoter, develop inflammatory skin disease and a form of murine lupus. To investigate the pathogenesis of this syndrome, we generated female IFN-γ transgenic mice congenitally deficient in either αβ or γδ T cells. TCRδ−/− transgenics continued to produce antinuclear autoantibodies and to develop severe kidney lesions. In contrast, TCRβ−/− IFN-γ transgenic mice failed to produce antinucleosome, anti-dsDNA, or antihistone autoantibodies, and kidney disease was abolished. Both αβ- and γδ-deficient transgenics continued to develop IFN-γ-associated skin disease, lymphadenopathy, and splenomegaly. The data show that the autoantibody-mediated pathology of murine lupus in IFN-γ transgenic mice is completely αβ T cell dependent and that γδ T cells cannot drive autoantibody production. These results imply that production of antinuclear autoantibodies in IFN-γ transgenic animals is Ag driven, and we identified clusters of apoptotic cells in the epidermis of the mice as a possible source of self Ags. Our findings emphasize the relevance of this murine lupus model to the human disease.
HLA-B15 Peptide Ligands Are Preferentially Anchored at Their C Termini
Authors: Prilliman Kiley R. , Jackson Kenneth W. , Lindsey Mark , Wang Jihua , Crawford David , Hildebrand William H. 
Published: December 2022

AbstractTherapies to elicit protective CTL require the selection of pathogen- and tumor-derived peptide ligands for presentation by MHC class I molecules. Edman sequencing of class I peptide pools generates “motifs” that indicate that nonameric ligands bearing conserved position 2 (P2) and P9 anchors provide the optimal search parameters for selecting immunogenic epitopes. To determine how well a motif represents its individual constituents, we used a hollow-fiber peptide production scheme followed by the mapping of endogenously processed class I peptide ligands through reverse-phase HPLC and mass spectrometry. Systematically mapping and characterizing ligands from B*1508, B*1501, B*1503, and B*1510 demonstrate that the peptides bound by these B15 allotypes i) vary in length from 7 to 12 residues, and ii) are more conserved at their C termini than their N-proximal P2 anchors. Comparative peptide mapping of these B15 allotypes further pinpoints endogenously processed ligands that bind to the allotypes B*1508, B*1501, and B*1503, but not B*1510. Overlapping peptide ligands are successful in binding to B*1501, B*1503, and B*1508 because these B15 allotypes share identical C-terminal anchoring pockets whereas B*1510 is divergent in the C-terminal pocket. Therefore, endogenous peptide loading into the B15 allotypes requires that a conserved C terminus be anchored in the appropriate specificity pocket while N-proximal anchors are more flexible in their location and sequence. Queries for overlapping and allele-specific peptide ligands may thus be contingent on a conserved C-terminal anchor.
Generation of Recombinant Fragments of CD11b Expressing the Functional β-Glucan-Binding Lectin Site of CR3 (CD11b/CD18)
Authors: Xia Yu , Ross Gordon D. 
Published: December 2022

AbstractCR3 (Mac-1; αMβ2 integrin) functions as both a receptor for the opsonic iC3b fragment of C3 triggering phagocytosis or cytotoxicity and an adhesion molecule mediating leukocyte diapedesis. Recent reports have suggested that a CR3 lectin site may be required for both cytotoxic responses and adhesion. Cytotoxic responses require dual recognition of iC3b via the I domain of CD11b and specific microbial surface polysaccharides (e.g., β-glucan) via a separate lectin site. Likewise, adhesion requires a lectin-dependent membrane complex between CR3 and CD87. To characterize the lectin site further, a recombinant baculovirus (rBv) system was developed that allowed high level expression of rCD11b on membranes and in the cytoplasm of Sf21 insect cells. Six rBv were generated that contained truncated cDNA encoding various CD11b domains. Immunoblotting of rBv-infected Sf21 cells showed that some native epitopes were expressed by five of six rCD11b fragments. Lectin activity of rCD11b proteins was evaluated by both flow cytometry with β-glucan-FITC and radioactive binding assays with [125I]β-glucan. Sf21 cells expressing rCD11b that included the C-terminal region, with or without the I-domain, exhibited lectin activity that was inhibited by unlabeled β-glucan or anti-CR3 mAbs. The smallest rCD11b fragment exhibiting lectin activity included the C-terminus and part of the divalent cation binding region. The β-glucan binding affinities of the three C-terminal region-containing rCD11bs expressed on Sf21 cell membranes were not significantly different from each other and were similar to that of neutrophil CR3. These data suggest that the lectin site may be located entirely within CD11b, although lectin site-dependent signaling through CD18 probably occurs with the heterodimer.
Differences Between IL-4- and IL-4 Receptor α-Deficient Mice in Chronic Leishmaniasis Reveal a Protective Role for IL-13 Receptor Signaling
Authors: Mohrs Markus , Ledermann Birgit , Köhler Gabriele , Dorfmüller Andreas , Gessner Andre , Brombacher Frank 
Published: December 2022

AbstractIL-4 receptor α-chain-deficient (IL-4Rα−/−) mice were generated by homologous and site-specific recombination, using the Cre/loxP system in BALB/c-derived embryonic stem cells. In vitro analysis of cells from these mice revealed impaired IL-4- and IL-13-mediated functions, demonstrating that the IL-4Rα-chain is an essential component of both the IL-4 and the IL-13 receptor. Whereas Leishmania major-infected BALB/c mice developed fatal progressive disease with type 2 Ab responses within 3 mo, both IL-4Rα−/− and IL-4−/− BALB/c mice contained infection with reduced footpad swelling, parasite load, moderate histopathology, and type 1 Ab responses during this time period. Conclusively, these results demonstrate an IL-4-dependent mechanism of susceptibility in BALB/c mice. Nevertheless, in contrast to mutant mice, infected C57BL/6 mice healed completely within 3 mo, indicating that additional factors are necessary for subsequent healing and elimination of the pathogen. During the further course of infection, IL-4Rα−/− mice developed progressive disease with massive footpad swelling. Lesions became ulcerative and necrotic with subsequent destruction of connective tissue and bones, as well as dissemination into organs and consequent mortality within the monitored 6 mo of chronic infection. In striking contrast, IL-4−/− mice maintained control of infection on a moderate level, but were unable to clear the pathogen. The distinct phenotypes of the BALB/c embryonic stem cell-derived IL-4−/− and IL-4Rα−/− mouse strains identify previously unsuspected mechanisms for maintaining host immunity to chronic infection with L. major, mediated by a functional IL-13 receptor.
Complex Requirements for Nascent and Memory Immunity in Pulmonary Histoplasmosis
Authors: Allendörfer Ruth , Brunner George D. , Deepe George S. 
Published: December 2022

AbstractThe presence of functional T cells is often required for successful resolution of infections with intracellular pathogens, yet the mechanisms by which they contribute to elimination of the invading pathogen in primary and secondary immunity are only partly understood. We report that increased mortality of naive α/β TCR+ or CD4+ T cell-depleted mice infected with the fungus Histoplasma capsulatum is associated with impairment of IFN-γ production. Upon secondary infection, mice concomitantly depleted of CD4+ and CD8+ cells exhibit decreased survival beyond day 25 of rechallenge, whereas elimination of either T cell subset or B cell deficiency does not result in accelerated mortality compared with controls. Remarkably, despite a decrease of H. capsulatum CFU in lungs of CD4+ plus CD8+-deficient mice, a progressive increase in spleen CFU is observed. The ability to control fungus growth in lungs is associated with vigorous TNF-α, but not IFN-γ, production by bronchoalveolar lavage cells. In contrast, spleen cells from CD4+ plus CD8+-deficient mice are unable to produce TNF-α. Thus, the cellular and molecular requirements for protective immunity vary between primary and secondary infection. Furthermore, in secondary histoplasmosis, a sharp contrast can be drawn between lungs and spleens in their reliance upon T cells to control fungal replication. The opposing activities of these organs can be ascribed in part to differential production of TNF-α.
Modulation of C3a Activity: Internalization of the Human C3a Receptor and its Inhibition by C5a
Authors: Settmacher Britta , Bock Daniel , Saad Henry , Gärtner Sören , Rheinheimer Claudia , Köhl Jörg , Bautsch Wilfried , Klos Andreas 
Published: December 2022

AbstractThe C3a receptor (C3aR) is expressed on most human peripheral blood leukocytes with the exception of resting lymphocytes, implying a much higher pathophysiological relevance of the anaphylatoxin C3a as a proinflammatory mediator than previously thought. The response to this complement split product must be tightly regulated in situations with sustained complement activation to avoid deleterious effects caused by overactivated inflammatory cells. Receptor internalization, an important control mechanism described for G protein-coupled receptors, was investigated. Using rabbit polyclonal anti-serum directed against the C3aR second extracellular loop, a flow cytometry-based receptor internalization assay was developed. Within minutes of C3a addition to human granulocytes, C3aR almost completely disappeared from the cell surface. C3aR internalization could also be induced by PMA, an activator of protein kinase C. Similarly, monocytes, the human mast cell line HMC-1, and differentiated monocyte/macrophage-like U937-cells exhibited rapid agonist-dependent receptor internalization. Neither C5a nor FMLP stimulated any cross-internalization of the C3aR. On the contrary, costimulation of granulocytes with C5a, but not FMLP, drastically decreased C3aR internalization. This effect could be blocked by a C5aR-neutralizing mAb. HEK293-cells transfected with the C3aR, with or without Gα16, a pertussis toxin-resistant G protein α subunit required for C3aR signal transduction in these cells, did not exhibit agonist-dependent C3aR internalization. Additionally, preincubation with pertussis toxin had no effect on C3a-induced internalization on PMNs. C3aR internalization is a rapid negative control mechanism and is influenced by the C5aR pathway.
Regulation of IFN Consensus Sequence Binding Protein Expression in Murine Macrophages
Authors: Kantakamalakul Wannee , Politis Alexander D. , Marecki Sylvia , Sullivan Teri , Ozato Keiko , Fenton Matthew J. , Vogel Stefanie N. 
Published: December 2022

AbstractRecent work has demonstrated that the transcription factor, IFN consensus sequence binding protein (ICSBP), plays a critical role in the capacity of mice to control infection with Toxoplasma gondii and Leishmania major, agents that require highly activated macrophages for their elimination. In this report the regulation of ICSBP mRNA and protein were analyzed in murine macrophages stimulated with LPS and/or IFN-γ. Like induction of leishmaniacidal activity, LPS and IFN-γ synergize to induce ICSBP mRNA and protein. Deletion analysis of the ICSBP promoter identified regions that were IFN-γ responsive, regions that mediate the ability of LPS and IFN-γ to activate this promoter synergistically, as well as regions that normally repress ICSBP transcription. Finally, exogenous expression of ICSBP, found in previous studies to down-regulate MHC I gene expression, failed to repress basal or IFN-γ-induced ICSBP transcription. This demonstrates that ICSBP can selectively suppress the expression of IFN-responsive genes. These findings extend in a significant way our understanding of the regulation of ICSBP by LPS and IFN-γ and provide important clues as to its role in macrophage activation.
Biased TCR Repertoire in HIV-1-Infected Patients Due to Clonal Expansion of HIV-1-Reverse Transcriptase-Specific CTL Clones
Authors: Kolowos W. , Schmitt M. , Herrman M. , Harrer E. , Löw P. , Kalden J. R. , Harrer T. 
Published: December 2022

AbstractTo study whether an expansion of HIV-1-specific CTL is contributing to the skewed TCR repertoire in HIV-1-infection, we characterized the TCR usage of CTL clones specific for a conserved epitope in HIV-1 reverse transcriptase (RT/476-484). CTL clones from three HIV-1-infected patients displayed highly similar TCR usage and used the identical Vβ6.1 and Vα2.5 gene segments. CTL clones from two patients showed a very high degree of similarity within the TCR complementarity-determining region-3 (CDR-3). In accordance with the similar molecular structure, all three CTL clones also exhibited a similar functional activity with regard to recognition of variant peptides and cytokine secretion pattern. In one subject clonal expansion of a single CTL specificity could be shown over a 10-mo period. TCR spectratyping of PBMC from two patients revealed a marked expansion of CDR-3 segments of a certain length within the Vβ6-family. Sequence analysis of these CDR-3 yielded sequences identical to the RT/476-484-specific CTL previously isolated from the same patients. This analysis demonstrates that clonal expansion of HIV-1-specific CTL is contributing to the skewed TCR repertoire in HIV-1-infected patients.
Characterization of Paired Ig-Like Receptors in Rats
Authors: Dennis Glynn , Stephan Robert P. , Kubagawa Hiromi , Cooper Max D. 
Published: December 2022

AbstractTo explore the phylogenetic history of the murine paired Ig-like receptors of activating (PIR-A) and inhibitory (PIR-B) types, we isolated PIR homologues from a rat splenocyte cDNA library. The rat (ra) PIR-A and raPIR-B cDNA sequences predict transmembrane proteins with six highly conserved extracellular Ig-like domains and distinctive membrane proximal, transmembrane, and cytoplasmic regions. The raPIR-B cytoplasmic region contains prototypic inhibitory motifs, whereas raPIR-A features a charged transmembrane region and a short cytoplasmic tail. Southern blot analysis predicts the presence of multiple Pira genes and a single Pirb gene in the rat genome. Although raPIR-A and raPIR-B are coordinately expressed by myeloid cells, analysis of mRNA detected unpaired expression of raPIR-A by B cells and raPIR-B by NK cells. Collectively, these findings indicate that the structural hallmarks of the Pir gene family are conserved in rats and mice, yet suggest divergence of PIR regulatory elements during rodent speciation.
T Suppressor Lymphocytes Inhibit NF-κB-Mediated Transcription of CD86 Gene in APC
Authors: Li Jianfeng , Liu Zhuoru , Jiang Shuiping , Cortesini Raffaello , Lederman Seth , Suciu-Foca Nicole 
Published: December 2022

AbstractCD8+CD28− human T suppressor cells (Ts) act on APC, inhibiting their ability to elicit Th activation and proliferation. This effect is due to inhibition of the CD40 pathway which normally leads to CD80 and CD86 up-regulation. To determine whether Ts inhibit expression of B7 molecules by blocking transcription, we cloned and characterized the CD86 promoter. Mutational analysis revealed that Ts inhibit transcription driven by the CD86 promoter. The NF-κB binding site, at −612 of the CD86 promoter, is essential for Th-induced transcription. In cultures containing Th and Ts, Ts inhibit Th-induced NF-κB activation in APC. Together, these findings indicate that Ts inhibition of NF-κB activation in APC is a means by which they regulate the activation and proliferation of Th.
Th1 and Th2 Deviation of Myelin-Autoreactive T Cells by Altered Peptide Ligands Is Associated with Reciprocal Regulation of Lck, Fyn, and ZAP-70
Authors: Singh Rana A. K. , Zang Ying C. Q. , Shrivastava Anju , Hong Jian , Wang George T. , Li Sufang , Tejada-Simon Maria V. , Kozovska Milena , Rivera Victor M. , Zhang Jingwu Z. 
Published: December 2022

AbstractTh0 clones recognizing an immunodominant peptide of myelin basic protein (residues 83–99) were derived from patients with multiple sclerosis. We demonstrate that analogue peptides with alanine substitution at Val86 and His88 had a unique partial agonistic property in inducing Th0 →Th1 and Th0 →Th2 deviation of the myelin basic protein-reactive T cell clones, respectively. Th0 to Th1 deviation induced by peptide 86V→A correlated with up-regulation of Fyn and ZAP-70 kinase activities. Conversely, Th0 to Th2 deviation induced by peptide 88H→A was associated with complete failure to activate Fyn and ZAP-70 kinases. The observed Th1 and Th2 shift also correlated, to a lesser extent, with Lck kinase activity that was down-regulated with Th1 deviation and increased with Th2 deviation in some T cell clones. We demonstrated that the Th1 and Th2 shift induced by the analogue peptides was a reversible process, as the T cell clones previously exposed to either 86V→A or 88H→A peptide could revert to an opposite phenotype when rechallenged reciprocally with a different analogue peptide. The study has important implications in our understanding of regulation of TCR-associated tyrosine kinases by altered peptide ligands and its role in cytokine regulation of autoreactive T cells.
Extracellular Signal-Related Kinase (ERK) and p38 Mitogen-Activated Protein (MAP) Kinases Differentially Regulate the Lipopolysaccharide-Mediated Induction of Inducible Nitric Oxide Synthase and IL-12 in Macrophages:LeishmaniaPhosphoglycans Subvert Macrophage IL-12 Production by Targeting ERK MAP Kinase
Authors: Feng Gui-Jie , Goodridge Helen S. , Harnett Margaret M. , Wei Xiao-Qing , Nikolaev Andrei V. , Higson Adrian P. , Liew Foo-Y. 
Published: December 2022

AbstractMacrophage activation by cytokines or microbial products such as LPS results in the induction and release of several key immune effector molecules including NO and IL-12. These have been shown to play crucial roles in the development of immunity to intracellular pathogens such as Leishmania. The molecular mechanisms underlying the induction of these effector molecules are not fully understood. We now show that the extracellular signal-related kinase (ERK) and p38 mitogen-activated protein (MAP) kinases play differential roles in the regulation of LPS-stimulated inducible NO synthase and IL-12 gene expression. In macrophages, LPS stimulates the simultaneous activation of all three classes of MAP kinases, ERK, c-jun N-terminal kinase, and p38, albeit with differential activation kinetics. However, studies using inhibitors selective for ERK (PD98059) and p38 (SB203580) show that while p38 plays an essential role in the induction of inducible NO synthase, ERK MAP kinases play only a minor role in promoting NO generation. In contrast, while p38 promotes induction of IL-12 (p40) mRNA, ERK activation suppresses LPS-mediated IL-12 transcription. The biological relevance of these regulatory signals is demonstrated by our finding that Leishmania lipophosphoglycans, which promote parasite survival, act by stimulating ERK MAP kinase to inhibit macrophage IL-12 production. Thus, as ERK and p38 MAP kinases differentially regulate the induction of the macrophage effector molecules, inducible NO synthase and IL-12, these kinases are potential targets not only for the development of novel strategies to combat intracellular pathogens but also for therapeutic immunomodulation.
Role of Immunoregulatory Donor T Cells in Suppression of Graft-Versus-Host Disease Following Donor Leukocyte Infusion Therapy
Authors: Johnson Bryon D. , Becker Emily E. , LaBelle James L. , Truitt Robert L. 
Published: December 2022

AbstractIn murine models of allogeneic bone marrow transplantation (BMT), MHC-mismatched recipients given a delayed infusion of donor leukocytes (DLI) at 21 days posttransplant develop significant GVHD whereas MHC-matched recipients do not. The current study was initially designed to test the hypothesis that small numbers of T cells in the MHC-mismatched donor bone marrow (BM) graft exacerbated graft-vs-host disease (GVHD) when DLI was administered at 21 days after BMT. Ex vivo depletion of Thy1+ cells from the donor BM had no impact on the severity of GVHD after DLI. However, depletion of donor T cells in vivo with a Thy1 allele-specific mAb given after BMT resulted in significantly more severe GVHD after DLI. Similar results were obtained in a MHC-matched model of allogeneic BMT, indicating that this was a general phenomenon and not model dependent. These results indicated that a population of donor-derived Thy1+ cells suppressed graft-vs-host reactivity after DLI. Results of experiments with thymectomized recipients demonstrated that an intact thymus was required for generation of the immunoregulatory donor cells. Experiments using TCR β-chain knockout mice as BM donors indicated that the immunosuppressive Thy1+ cells coexpressed αβTCR heterodimers. Similar experiments with CD4 and CD8 knockout donor BM suggested that the immunoregulatory Thy1+αβTCR+ cells consisted of two subpopulations: a CD4+CD8− subpopulation and a CD4−CD8− subpopulation. Together, these results show that thymus-derived, Thy1+αβTCR+ donor cells generated early after allogeneic BMT suppress the graft-vs-host reactivity of T cells given as DLI. These cells may mediate dominant peripheral tolerance after allogeneic BMT.