bluuebunny/crossref_metadata_2025_split huggingface dataset:
Functional Characterization of Human Cysteinyl Leukotriene 1 Receptor Gene Structure
Authors: Woszczek Grzegorz , Pawliczak Rafal , Qi Hai-Yan , Nagineni Sahrudaya , Alsaaty Sura , Logun Carolea , Shelhamer James H. 
Published: April 2014

Abstract
The 5-lipoxygenase pathway has been strongly implicated in the pathogenesis of chronic inflammatory disorders, such as bronchial asthma and atherosclerosis. Cysteinyl leukotrienes (cysLTs), 5-lipoxygenase pathway products, are recognized now not only as important factors in asthmatic inflammation, but also as mediators of cell trafficking and innate immune responses. To study a role of cysLTs in inflammatory reactions we have characterized the gene structure of human cysteinyl leukotriene receptor type I (cysLT1R). The cysLT1R gene consists of 5 exons that are variably spliced and a single promoter region with multiple transcription start sites. Four different cysLT1R transcripts were identified. RT-PCR showed dominant and wide expression of the transcript I, containing exons 1, 4, and 5, with the strongest presence in blood leukocytes, spleen, thymus, lung, and heart. The expression of cysLT1R is functionally regulated at the transcriptional level by IL-4 through a STAT6 response element localized to the proximal cysLT1R promoter region. IL-4 stimulation increased cysLT1R mRNA (real-time PCR) and surface protein expression (flow cytometry) in a time-dependent fashion. CysLTs (LTD4 and LTC4) induced an increased production of a potent monocyte chemoattractant CCL2 (MCP-1) in IL-4-primed THP-1 cells in a dose-dependent manner. This effect was effectively inhibited by the cysLT1R-selective antagonist MK571 in a dose-dependent manner and only partially by a nonselective cysLT1R/cysLT2R inhibitor BAY-u9773, implying a cysLT1R-mediated mechanism. Thus, cysLTs signaling through cysLT1R might contribute to inflammatory reactions by cooperating with IL-4 in enhanced CCL2 production in human monocytic cells.
Hypermutation at A-T Base Pairs: The A Nucleotide Replacement Spectrum Is Affected by Adjacent Nucleotides and There Is No Reverse Complementarity of Sequences Flanking Mutated A and T Nucleotides
Authors: Spencer Jo , Dunn-Walters Deborah K. 
Published: April 2014

Abstract
Hypermutation is thought to be a two-phase process. The first phase is via the action of activation-induced cytidine deaminase (AID), which deaminates C nucleotides in WRC motifs. This results in the RGYW/WRCY hot spot motifs for mutation from G and C observed in vivo. The resemblance between the hot spot for C mutations and the reverse complement of that for G mutations implies a process acting equally on both strands of DNA. The second phase of hypermutation generates mutations from A and T and exhibits strand bias, with more mutations from A than T. Although this does not concur with the idea of one mechanism acting equally on both strands, it has been suggested that the AT mutator also has a reversible motif; WA/TW. We show here that the motifs surrounding the different substitutions from A vary significantly; there is no single targeting motif for all A mutations. Sequence preferences associated with mutations from A more likely reflect an influence of adjacent nucleotides over what the A mutates “to.” This influence tends toward “like” replacements: Purines (A or G) in the 5′ position bias toward replacement by another purine (G), whereas replacement with pyrimidines (C or T) is more likely if the preceding base is also a pyrimidine. There is no reverse complementarity in these observations, in that similar influences of nucleotides adjacent to T are not seen. Hence, WA and TW should not be considered as reverse complement hot spot motifs for A and T mutations.
Sequence Motifs in IL-4Rα Mediating Cell-Cycle Progression of Primary Lymphocytes
Authors: Stephenson Linda M. , Park Do-Sim , Mora Ana L. , Goenka Shreevrat , Boothby Mark 
Published: April 2014

Abstract
IL-4 signaling through the IL-4Rα chain regulates the development and proliferation of the Th2 lineage of effector CD4+ T cells. Analyses of the IL-4R in factor-dependent cell lines led to the development of two apparently conflicting models of the primary structural determinants of IL-4R-mediated proliferative signaling. In one model, proliferation was dependent on the first conserved tyrosine in the cytoplasmic tail (Y1), while in the second, proliferation was independent of cytoplasmic tyrosines. We found that in activated primary T cells, mutation of only the Y1 residue resulted in a modest decrease in IL-4-induced S phase entry, a further decrease in cell-cycle completion, and a complete failure of IL-4 to induce p70S6 kinase phosphorylation. Consistent with a role for the PI3K/mammalian target of rapamycin pathway in mediating cytokine acceleration of G2/M transit, pretreatment of activated T cells with rapamycin resulted in only a modest decrease in IL-4-induced S phase entry, but a total block of cell-cycle completion. Strikingly, IL-4Rα chains that lacked all cytoplasmic tyrosines were competent to signal for STAT5 phosphorylation, mediated efficient S phase entry, and promoted cell-cycle progression. The ability of tyrosine-deficient IL-4Rs to mediate proliferative signaling and STAT phosphorylation was absolutely dependent on the presence of an intact ID-1 region. These findings show that IL-4Rα lacking cytoplasmic tyrosine residues is competent to induce ID-1-dependent proliferation, and indicate that IL-4 can promote G2/M progression via activation of the mammalian target of rapamycin pathway initiated at the Y1 residue.
Allele-Specific Regulation of TCRβ Variable Gene Segment Chromatin Structure
Authors: Jackson Annette M. , Krangel Michael S. 
Published: April 2014

Abstract
Allelic exclusion of the murine Tcrb locus is imposed at the level of recombination and restricts each cell to produce one functional VDJβ rearrangement. Allelic exclusion is achieved through asynchronous Vβ to DJβ recombination as well as feedback inhibition that terminates recombination once a functional rearrangement has occurred. Because the accessibility of Vβ gene segment chromatin is diminished as thymocytes undergo allelic exclusion at the CD4−CD8− (double-negative) to CD4+CD8+ (double-positive) transition, chromatin regulation was thought to be an important component of the feedback inhibition process. However, previous studies of chromatin regulation addressed the status of Tcrb alleles using genetic models in which both alleles remained in a germline configuration. Under physiological conditions, developing thymocytes would undergo Vβ to DJβ recombination on one or both alleles before the enforcement of feedback. On rearranged alleles, Vβ gene segments that in germline configuration are regulated independently of the Tcrb enhancer are now brought into its proximity. We show in this study that in contrast to Vβ segments on a nonrearranged allele, those situated upstream of a functionally rearranged Vβ segment are contained in active chromatin as judged by histone H3 acetylation, histone H3 lysine 4 (K4) methylation, and germline transcription. Nevertheless, these Vβ gene segments remain refractory to recombination in double-positive thymocytes. These results suggest that a unique feedback mechanism may operate independent of chromatin structure to inhibit Vβ to DJβ recombination after the double-negative stage of thymocyte development.
Splenic Marginal Zone Dendritic Cells Mediate the Cholera Toxin Adjuvant Effect: Dependence on the ADP-Ribosyltransferase Activity of the Holotoxin
Authors: Grdic Dubravka , Ekman Lena , Schön Karin , Lindgren Kristina , Mattsson Johan , Magnusson Karl-Eric , Ricciardi-Castagnoli Paola , Lycke Nils 
Published: April 2014

Abstract
The in vivo mechanisms of action of most vaccine adjuvants are poorly understood. In this study, we present data in mice that reveal a series of critical interactions between the cholera toxin (CT) adjuvant and the dendritic cells (DC) of the splenic marginal zone (MZ) that lead to effective priming of an immune response. For the first time, we have followed adjuvant targeting of MZ DC in vivo. We used CT-conjugated OVA and found that the Ag selectively accumulated in MZ DC following i.v. injections. The uptake of Ag into DC was GM1 ganglioside receptor dependent and mediated by the B subunit of CT (CTB). The targeted MZ DC were quite unique in their phenotype: CD11c+, CD8α−, CD11b−, B220−, and expressing intermediate or low levels of MHC class II and DEC205. Whereas CTB only delivered the Ag to MZ DC, the ADP-ribosyltransferase activity of CT was required for the maturation and migration of DC to the T cell zone, where these cells distinctly up-regulated CD86, but not CD80. This interaction appeared to instruct Ag-specific CD4+ T cells to move into the B cell follicle and strongly support germinal center formations. These events may explain why CT-conjugated Ag is substantially more immunogenic than Ag admixed with soluble CT and why CTB-conjugated Ag can tolerize immune responses when given orally or at other mucosal sites.
Human High Mobility Group Box Transcription Factor 1 Affects Thymocyte Development and Transgene Variegation
Authors: Sekkali Belaïd , Szabat Ewa , Ktistaki Eleni , Tolaini Mauro , Roderick Kathleen , Harker Nicky , Patel Amisha , Williams Keith , Norton Trisha , Kioussis Dimitris 
Published: April 2014

Abstract
It has been shown previously that a human CD2 (hCD2) disabled locus control region (LCR) transgene is unable to establish an open chromatin configuration in all the T cells, and this leads to position effect variegation of the transgene. In this study we show that thymus-specific overexpression of human high mobility group box transcription factor 1 (HBP1), a transcription factor that binds a specific sequence within the hCD2 LCR, affects thymus cellularity as well as the number of CD8+ thymocytes in two independent transgenic mouse lines and increases the proportion of T cells that fully activate the transgenic locus in hCD2 variegating mice in a sequence-specific dependent manner. This finding suggests that overexpression of HBP1 can affect lineage commitment and can relieve the suppressive influence of heterochromatin, allowing thymocytes to express the variegating target locus more efficiently. These effects could be the result of direct HBP1 action on LCR activity. Alternatively, the extra HBP1 molecules may sequester repressive elements away from the LCR, thus allowing transcription permissive states to form on the transgene locus.
Unusually High Frequency MHC Class I Alleles in Mauritian Origin Cynomolgus Macaques
Authors: Krebs Kendall C. , Jin ZheYuan , Rudersdorf Richard , Hughes Austin L. , O’Connor David H. 
Published: April 2014

Abstract
Acute shortages of Indian origin Rhesus macaques significantly hinder HIV/AIDS research. Cellular immune responses are particularly difficult to study because only a subset of animals possess MHC class I (MHC I) alleles with defined peptide-binding specificities. To expand the pool of nonhuman primates suitable for studies of cellular immunity, we defined 66 MHC I alleles in Cynomolgus macaques (Macaca fascicularis) of Chinese, Vietnamese, and Mauritian origin. Most MHC I alleles were found only in animals from a single geographic origin, suggesting that Cynomolgus macaques from different origins are not interchangeable in studies of cellular immunity. Animals from Mauritius may be particularly valuable because &gt;50% of these Cynomolgus macaques share the MHC class I allele combination Mafa-B*430101, Mafa-B*440101, and Mafa-B*460101. The increased MHC I allele sharing of Mauritian origin Cynomolgus macaques may dramatically reduce the overall number of animals needed to study cellular immune responses in nonhuman primates while simultaneously reducing the confounding effects of genetic heterogeneity in HIV/AIDS research.
The Murine Family of Gut-Restricted Class Ib MHC Includes Alternatively Spliced Isoforms of the Proposed HLA-G Homolog, “Blastocyst MHC”
Authors: Guidry Paula A. , Stroynowski Iwona 
Published: April 2014

Abstract
The gastrointestinal tract is populated by a multitude of specialized immune cells endowed with receptors for classical (class Ia) and nonclassical (class Ib) MHC proteins. To identify class I products that engage these receptors and impact immunity/tolerance, we studied gut-transcribed class Ib loci and their polymorphism in inbred, outbred, and wild-derived mice. Intestinal tissues enriched in epithelial cells contained abundant transcripts of ubiquitously expressed and preferentially gut-restricted Q and T class I loci. The latter category included the “blastocyst Mhc” gene, H2-Bl, and its putative paralog, Tw5. Expression of H2-Bl was previously detected only at the maternal/fetal interface, where it was proposed to induce immune tolerance via interactions with CD94/NKG2A receptors. Analysis of coding region polymorphism performed here revealed two major alleles of H2-Bl with conserved residues at positions critical for class I protein folding and peptide binding. Both divergent alleles are maintained in outbred and wild mice under selection for fecundity and pathogen resistance. Surprisingly, we found that alternative splicing of H2-Bl mRNA in gut tissues is prevalent and allele-specific. It leads to strain-dependent expression of diverse repertoires of canonical and noncanonical transcripts that may give rise to distinct ligands for intestinal NK cell, T cell, and/or intraepithelial lymphocyte receptors.
Increase in Phagocytosis after Geldanamycin Treatment or Heat Shock: Role of Heat Shock Proteins
Authors: Vega Virginia L. , De Maio Antonio 
Published: April 2014

Abstract
The response to injury is activated at the systemic and cellular levels. At the systemic level, phagocytosis plays a key role in controlling infections and clearing necrotic and apoptotic cells. The expression of heat shock proteins (Hsp), which is a well-conserved process, is a major component of cellular response to stress. This study investigated the relationship between Hsps and phagocytosis. An increase in the phagocytosis of opsonized bacteria particles and latex beads was observed upon incubation of murine macrophages with geldanamycin (GA), a specific inhibitor of the Hsp90 family of proteins. The effect of GA on phagocytosis was blocked by coincubation with inhibitors of transcription (actinomycin D) or translation (cycloheximide), suggesting that gene expression was required. Because expression of Hsps has been observed after GA treatment, the effect of heat shock on phagocytosis was investigated. Similar to GA treatment, heat shock resulted in an actinomycin D-sensitive elevation of phagocytosis, which suggests that Hsps are involved. The increase in phagocytosis after GA treatment was not due to increased binding of opsonized particles to their respective receptors on the macrophage surface or to elevated oxidative stress. However, it was correlated with a rapid polymerization of actin in proximity to the plasma membrane. These results suggest that Hsps play a role in the modulation of the phagocytic process, which is part of the stress response.
IL-12 Is Required for Induction but Not Maintenance of Protective, Memory Responses to Blastomyces dermatitidis: Implications for Vaccine Development in Immune-Deficient Hosts
Authors: Wüthrich Marcel , Warner Tom , Klein Bruce S. 
Published: April 2014

Abstract
Cellular immunity mediated by T lymphocytes, in particular CD4+ and CD8+ type 1 (T1) cells, is the main defense against pathogenic fungi. IL-12 initiates T1 cell development and cell-mediated immunity, but it is unclear whether IL-12 contributes to the maintenance of an antifungal T1 response. In this study, we addressed the role of IL-12 for vaccine-induced memory T cell development against experimental pulmonary blastomycosis. CD4+ T cells absolutely required IL-12 to control a live genetically engineered attenuated strain of Blastomyces dermatitidis given s.c. as a vaccine, whereas CD8+ T cells were significantly less dependent on IL-12. Despite differential dependency of T cell subsets on IL-12 during vaccination, neither subset acquired memory immunity in the absence of IL-12. In contrast, adoptive transfer of immune CD4 T cells from wild-type mice into IL-12−/− mice showed that CD4+ T1 memory cells sustained a T1 cytokine profile and remained protective over a period of 6 mo posttransfer. Similarly, memory CD8 cells elicited in IL-12−/− mice with killed yeast and transient rIL-12 treatment (during vaccination) remained durable and protective after animals were rested for 3 mo. In conclusion, these studies demonstrate that once CD4 and CD8 cells have acquired a protective T1 phenotype they no longer require the presence of IL-12 to maintain antifungal protective memory.
Cloning of the Gene Encoding a Protective Mycobacterium tuberculosis Secreted Protein Detected In Vivo during the Initial Phases of the Infectious Process
Authors: Mukherjee Sandeep , Kashino Suely S. , Zhang Yanni , Daifalla Nada , Rodrigues Virmondes , Reed Steven G. , Campos-Neto Antonio 
Published: April 2014

Abstract
The existence of therapeutic agents and the bacille Calmette-Guérin (BCG) vaccine have not significantly affected the current tuberculosis pandemic. BCG vaccine protects against serious pediatric forms of tuberculosis but not against adult pulmonary tuberculosis, the most common and contagious form of the disease. Several vaccine candidates, including Mycobacterium tuberculosis recombinant proteins formulated in newer adjuvants or delivered in bacterial plasmid DNA have recently been described. An attractive source of vaccine candidates has been M. tuberculosis Ags present in culture supernatants of the initial phases of the bacterial growth in vitro. In this study we describe an Ag discovery approach to select for such Ags produced in vivo during the initial phases of the infection. We combined RP-HPLC and mass spectrometry to identify secreted or shed M. tuberculosis proteins eliminated in animal urine within 14 days after the infection. A peptide containing sequence homology with a hypothetical M. tuberculosis protein was identified and the recombinant protein produced in Escherichia coli. The protein was recognized by Ab (IgG2a and IgG1) and T cells (Th1) of mice infected with M. tuberculosis and by lymphoid cells from healthy donors who had a positive purified protein derivative skin test but not from tuberculosis patients. Moreover, this Ag induced protection in mice against M. tuberculosis at levels comparable to protection induced by BCG vaccine. These results validate the Ag discovery approach of M. tuberculosis proteins secreted or shed in vivo during the early phases of the infection and open new possibilities for the development of potential vaccine candidates or of markers of active mycobacterial multiplication and therefore active disease.
Mycobacterium bovis BCG Attenuates Surface Expression of Mature Class II Molecules through IL-10-Dependent Inhibition of Cathepsin S
Authors: Sendide Khalid , Deghmane Ala-Eddine , Pechkovsky Dmitri , Av-Gay Yossef , Talal Amina , Hmama Zakaria 
Published: April 2014

Abstract
We have previously shown that macrophage infection with Mycobacterium tuberculosis and M. bovis bacillus Calmette-Guérin (BCG) partially inhibits MHC class II surface expression in response to IFN-γ. The present study examined the nature of class II molecules that do in fact reach the surface of infected cells. Immunostaining with specific Abs that discriminate between mature and immature class II populations showed a predominance of invariant chain (Ii)-associated class II molecules at the surface of BCG-infected cells suggesting that mycobacteria specifically block the surface export of peptide-loaded class II molecules. This phenotype was due to inhibition of IFN-γ-induced cathepsin S (Cat S) expression in infected cells and the subsequent intracellular accumulation of αβ class II dimers associated with the Cat S substrate Ii p10 fragment. In contrast, infection with BCG was shown to induce secretion of IL-10, and addition of blocking anti-IL-10 Abs to cell cultures restored both expression of active Cat S and export of mature class II molecules to the surface of infected cells. Consistent with these findings, expression of mature class II molecules was also restored in cells infected with BCG and transfected with active recombinant Cat S. Thus, M. bovis BCG exploits IL-10 induction to inhibit Cat S-dependent processing of Ii in human macrophages. This effect results in inhibition of peptide loading of class II molecules and in reduced presentation of mycobacterial peptides to CD4+ T cells. This ability may represent an effective mycobacterial strategy for eluding immune surveillance and persisting in the host.
Characterization of a Novel Chemokine-Containing Storage Granule in Endothelial Cells: Evidence for Preferential Exocytosis Mediated by Protein Kinase A and Diacylglycerol
Authors: Øynebråten Inger , Barois Nicolas , Hagelsteen Kathrine , Johansen Finn-Eirik , Bakke Oddmund , Haraldsen Guttorm 
Published: April 2014

Abstract
We have recently shown that several proinflammatory chemokines can be stored in secretory granules of endothelial cells (ECs). Subsequent regulated exocytosis of such chemokines may then enable rapid recruitment of leukocytes to inflammatory sites. Although IL-8/CXCL8 and eotaxin-3/CCL26 are sorted to the rod-shaped Weibel-Palade body (WPB), we found that GROα/CXCL1 and MCP-1/CCL2 reside in small granules that, similarly to the WPB, respond to secretagogue stimuli. In the present study, we report that GROα and MCP-1 colocalized in 50- to 100-nm granules, which occur throughout the cytoplasm and at the cell cortex. Immunofluorescence confocal microscopy revealed no colocalization with multimerin or tissue plasminogen activator, i.e., proteins that are released from small granules of ECs by regulated exocytosis. Moreover, the GROα/MCP-1-containing granules were Rab27-negative, contrasting the Rab27-positive, WPB. The secretagogues PMA, histamine, and forskolin triggered distinct dose and time-dependent responses of GROα release. Furthermore, GROα release was more sensitive than IL-8 release to inhibitors and activators of PKA and PKC but not to an activator of Epac, a cAMP-regulated GTPase exchange factor, indicating that GROα release is regulated by molecular adaptors different from those regulating exocytosis of the WPB. On the basis of these findings, we designated the GROα/MCP-1-containing compartment the type 2 granule of regulated secretion in ECs, considering the WPB the type 1 compartment. In conclusion, we propose that the GROα/MCP-1-containing type 2 granule shows preferential responsiveness to important mediators of EC activation, pointing to the existence of selective agonists that would allow differential release of selected chemokines.