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111.
Miyahira Y Akiba H Katae M Kubota K Kobayashi S Takeuchi T García-Sastre A Fukuchi Y Okumura K Yagita H Aoki T 《Journal of immunology (Baltimore, Md. : 1950)》2003,171(12):6344-6348
The ligand to receptor activator of NF-kappaB (RANK-L)/RANK interaction has been implicated in CD40 ligand/CD40-independent T cell priming by dendritic cells. In this report, we show that the coadministration of the RANK-L gene with a Trypanosoma cruzi gene markedly enhances the induction of Trypanosoma Ag-specific CD8(+) T cells and improves the DNA vaccine efficacy. A similarly potent adjuvant effect of the RANK-L gene on the induction of Ag-specific CD8(+) T cells was also observed when recombinant influenza virus expressing murine malaria Ag was used as an immunogen. In contrast, the coadministration of the CD40L gene was not effective in these systems. Our results demonstrated, for the first time, the potent immunostimulatory effect of the RANK-L gene to improve the CD8(+) T cell-mediated immunity against infectious agents. 相似文献
112.
Uemura Y Senju S Maenaka K Iwai LK Fujii S Tabata H Tsukamoto H Hirata S Chen YZ Nishimura Y 《Journal of immunology (Baltimore, Md. : 1950)》2003,170(2):947-960
Accumulating evidence indicates that recognition by TCRs is far more degenerate than formerly presumed. Cross-recognition of microbial Ags by autoreactive T cells is implicated in the development of autoimmunity, and elucidating the recognition nature of TCRs has great significance for revelation of the disease process. A major drawback of currently used means, including positional scanning synthetic combinatorial peptide libraries, to analyze diversity of epitopes recognized by certain TCRs is that the systematic detection of cross-recognized epitopes considering the combinatorial effect of amino acids within the epitope is difficult. We devised a novel method to resolve this issue and used it to analyze cross-recognition profiles of two glutamic acid decarboxylase 65-autoreactive CD4(+) T cell clones, established from type I diabetes patients. We generated a DNA-based randomized epitope library based on the original glutamic acid decarboxylase epitope using class II-associated invariant chain peptide-substituted invariant chains. The epitope library was composed of seven sublibraries, in which three successive residues within the epitope were randomized simultaneously. Analysis of agonistic epitopes indicates that recognition by both TCRs was significantly affected by combinations of amino acids in the antigenic peptide, although the degree of combinatorial effect differed between the two TCRs. Protein database searching based on the TCR recognition profile proved successful in identifying several microbial and self-protein-derived mimicry epitopes. Some of the identified mimicry epitopes were actually produced from recombinant microbial proteins by APCs to stimulate T cell clones. Our data demonstrate the importance of the combinatorial nature of amino acid residues of epitopes in molecular mimicry. 相似文献
113.
Conserved protein kinases encoded by herpesviruses and cellular protein kinase cdc2 target the same phosphorylation site in eukaryotic elongation factor 1delta 下载免费PDF全文
Kawaguchi Y Kato K Tanaka M Kanamori M Nishiyama Y Yamanashi Y 《Journal of virology》2003,77(4):2359-2368
Earlier studies have shown that translation elongation factor 1delta (EF-1delta) is hyperphosphorylated in various mammalian cells infected with representative alpha-, beta-, and gammaherpesviruses and that the modification is mediated by conserved viral protein kinases encoded by herpesviruses, including UL13 of herpes simplex virus type 1 (HSV-1), UL97 of human cytomegalovirus, and BGLF4 of Epstein-Barr virus (EBV). In the present study, we attempted to identify the site in EF-1delta associated with the hyperphosphorylation by the herpesvirus protein kinases. Our results are as follows: (i) not only in infected cells but also in uninfected cells, replacement of the serine residue at position 133 (Ser-133) of EF-1delta by alanine precluded the posttranslational processing of EF-1delta, which corresponds to the hyperphosphorylation. (ii) A purified chimeric protein consisting of maltose binding protein (MBP) fused to a domain of EF-1delta containing Ser-133 (MBP-EFWt) is specifically phosphorylated in in vitro kinase assays by purified recombinant UL13 fused to glutathione S-transferase (GST) expressed in the baculovirus system. In contrast, the level of phosphorylation by the recombinant UL13 of MBP-EFWt carrying an alanine replacement of Ser-133 (MBP-EFS133A) was greatly impaired. (iii) MBP-EFWt is also specifically phosphorylated in vitro by purified recombinant BGLF4 fused to GST expressed in the baculovirus system, and the level of phosphorylation of MBP-EFS133A by the recombinant BGLF4 was greatly reduced. (iv) The sequence flanking Ser-133 of EF-1delta completely matches the consensus phosphorylation site for a cellular protein kinase, cdc2, and in vitro kinase assays revealed that purified cdc2 phosphorylates Ser-133 of EF-1delta. (v) As observed with EF-1delta, the casein kinase II beta subunit (CKIIbeta) was specifically phosphorylated by UL13 in vitro, while the level of phosphorylation of CKIIbeta by UL13 was greatly diminished when a serine residue at position 209, which has been reported to be phosphorylated by cdc2, was replaced with alanine. These results indicate that the conserved protein kinases encoded by herpesviruses and a cellular protein kinase, cdc2, have the ability to target the same amino acid residues for phosphorylation. Our results raise the possibility that the viral protein kinases mimic cdc2 in infected cells. 相似文献
114.
Yahata Y Shirakata Y Tokumaru S Yamasaki K Sayama K Hanakawa Y Detmar M Hashimoto K 《The Journal of biological chemistry》2003,278(41):40026-40031
115.
Although the expression of the metastases-associated gene MTA1 correlates with tumor metastases, its role in regulating type IV collagenase expression is unknown. Enforced MTA1 expression in HT1080 cells reduced basal and 12-myristate 13-acetate-induced 92-kDa type IV collagenase (MMP-9) protein/mRNA levels. DNase I hypersensitivity and PstI accessibility assays revealed multiple regions of the MMP-9 promoter (-650/-450 and -120/+1), showing reduced hypersensitivity in the MTA1-expressing cells. Chromatin immunoprecipitation assays demonstrated MTA1 binding to the distal region, which spans several regulatory cis elements. Co-immunoprecipitation and chromatin immunoprecipitation assay experiments revealed histone deacetylase 2 (HDAC2)-MTA1 protein-protein interactions and the MTA1-dependent recruitment of HDAC2 to the distal MMP-9 promoter region, yielding diminished histone H3/H4 acetylation. However, HDAC2 binding and H3/H4 acetylation at the proximal MMP-9 region were unaffected by MTA1 expression. Furthermore, trichostatin treatment only partially relieved MTA1-repressed MMP-9 expression, indicating a HDAC-insensitive component possibly involv ing the nucleosome-remodeling Mi2 activity, which was recruited to the promoter by MTA1. In summary, (a) MMP-9 adds to a short list of MTA1-regulated genes, which so far only includes c-myc and pS2, and (b) MTA1 binds to the MMP-9 promoter, thereby repressing expression of this type IV collagenase via histone-dependent and independent mechanisms. 相似文献
116.
Phosphoenolpyruvate carboxylase (PEPC; EC 4.1.1.31) catalyzes the irreversible carboxylation of phosphoenolpyruvate (PEP) to form oxaloacetate and Pi using Mg2+ or Mn2+ as a cofactor. PEPC plays a key role in photosynthesis by C4 and Crassulacean acid metabolism plants, in addition to its many anaplerotic functions. Recently, three-dimensional structures of PEPC from Escherichia coli and the C4 plant maize (Zea mays) were elucidated by X-ray crystallographic analysis. These structures reveal an overall square arrangement of the four identical subunits, making up a "dimer-of-dimers" and an eight-stranded beta barrel structure. At the C-terminal region of the beta barrel, the Mn2+ and a PEP analog interact with catalytically essential residues, confirmed by site-directed mutagenesis studies. At about 20A from the beta barrel, an allosteric inhibitor (aspartate) was found to be tightly bound to down-regulate the activity of the E. coli enzyme. In the case of maize C4-PEPC, the putative binding site for an allosteric activator (glucose 6-phosphate) was also revealed. Detailed comparison of the various structures of E. coli PEPC in its inactive state with maize PEPC in its active state shows that the relative orientations of the two subunits in the basal "dimer" are different, implicating an allosteric transition. Dynamic movements were observed for several loops due to the binding of either an allosteric inhibitor, a metal cofactor, a PEP analog, or a sulfate anion, indicating the functional significance of these mobile loops in catalysis and regulation. Information derived from these three-dimensional structures, combined with related biochemical studies, has established models for the reaction mechanism and allosteric regulation of this important C-fixing enzyme. 相似文献
117.
Homocysteine induces vascular endothelial growth factor expression in differentiated THP-1 macrophages 总被引:10,自引:0,他引:10
Hyperhomocysteinemia has been reported to be an independent risk factor for atherosclerosis and atherothrombosis. However, the molecular mechanism by which hyperhomocysteinemia can lead to atherosclerosis and atherothrombosis has not been completely described. Vascular endothelial growth factor (VEGF) has been proposed to play an important role in the progression of atherosclerosis. In the present study, we hypothesized that hyperhomocysteinemia might be associated with VEGF expression in atherosclerotic lesions. We investigated VEGF mRNA expression and VEGF secretion by homocysteine (Hcy) in differentiated THP-1 macrophages. As a result, it has been revealed that VEGF mRNA was upregulated by Hcy in a dose- and time-dependent manner in THP-1 macrophages with the increase in VEGF secretion. Importantly, other sulfur compounds, such as methionine and cysteine, showed no effect on VEGF expression, indicating that homocysteine specifically induced VEGF. Our findings suggest that hyperhomocysteinemia could promote the development of atherosclerotic lesions through VEGF induction in macrophages. 相似文献
118.
Shimosato T Kitazawa H Katoh S Tomioka Y Karima R Ueha S Kawai Y Hishinuma T Matsushima K Saito T 《Biochimica et biophysica acta》2003,1627(1):56-61
Complementary DNA (cDNA) encoding swine Toll-like receptor 9 (sTLR9) was isolated from Peyer's patches (Pps) of gut-associated lymphoid tissue (GALT). The complete open reading frame (ORF) of sTLR9 contains 3093 bp coding deduced 1030 amino acid residues. The amino acid sequence of sTLR9 was characterized by a signal peptide followed by multiple leucine-rich repeats, a transmembrane sequence and a cytoplasmic domain homologous to that of the human interleukin-1 receptor (TIR). The sTLR9 showed a higher amino acid identity with humans (81.8%) and felis catus (86.7%) than mice (74.9%). The HEK293T cells transfected with pCXN2.1-FLAG DNA containing the sTLR9 cDNA were expressed sTLR9 as a membrane-bound molecules, which were reactive with anti-sTLR9 rabbit polyclonal antibody. Moreover, the transfectant was responsible for the CpG oligo DNA. sTLR9 was preferentially expressed in Pps and mesenteric lymph nodes (MLNs), and its degree was approximately three times higher than a spleen but weak in the other tissues by the real-time quantitative PCR analyses. The strong expression of sTLR9 in Pps and MLNs and its recognizing CpG DNA for human cell stimulant are shown first in this study, which may help in understanding the intestinal immune system mediated by a bacterial DNA through TLR9. 相似文献
119.
Siegel DH Ashton GH Penagos HG Lee JV Feiler HS Wilhelmsen KC South AP Smith FJ Prescott AR Wessagowit V Oyama N Akiyama M Al Aboud D Al Aboud K Al Githami A Al Hawsawi K Al Ismaily A Al-Suwaid R Atherton DJ Caputo R Fine JD Frieden IJ Fuchs E Haber RM Harada T Kitajima Y Mallory SB Ogawa H Sahin S Shimizu H Suga Y Tadini G Tsuchiya K Wiebe CB Wojnarowska F Zaghloul AB Hamada T Mallipeddi R Eady RA McLean WH McGrath JA Epstein EH 《American journal of human genetics》2003,73(1):174-187
Kindler syndrome is an autosomal recessive disorder characterized by neonatal blistering, sun sensitivity, atrophy, abnormal pigmentation, and fragility of the skin. Linkage and homozygosity analysis in an isolated Panamanian cohort and in additional inbred families mapped the gene to 20p12.3. Loss-of-function mutations were identified in the FLJ20116 gene (renamed “KIND1” [encoding kindlin-1]). Kindlin-1 is a human homolog of the Caenorhabditis elegans protein UNC-112, a membrane-associated structural/signaling protein that has been implicated in linking the actin cytoskeleton to the extracellular matrix (ECM). Thus, Kindler syndrome is, to our knowledge, the first skin fragility disorder caused by a defect in actin-ECM linkage, rather than keratin-ECM linkage. 相似文献
120.
DNA microarray analysis of human gingival fibroblasts from healthy and inflammatory gingival tissues 总被引:5,自引:0,他引:5
Wang PL Ohura K Fujii T Oido-Mori M Kowashi Y Kikuchi M Suetsugu Y Tanaka J 《Biochemical and biophysical research communications》2003,305(4):970-973
In the inflammatory gingival tissues of patients with periodontitis, cytokines such as interleukin (IL)-1 alpha, IL-1 beta, IL-6, IL-8, and tumor necrosis factor (TNF)-alpha have been detected. Gingival fibroblasts are the major constituents of gingival tissue. We recently demonstrated that lipopolysaccharide (LPS) from periodontopathic bacteria induces inflammatory reactions in various tissues via CD14 and/or Toll-like receptors (TLRs) in gingival tissues [Biochem. Biophys. Res. Commun. 273 (2000) 1161]. To confirm this, we examined the expression of IL-1 alpha, IL-1 beta, IL-6, IL-8, TNF-alpha, CD14, TLR2, and TLR4 in human gingival fibroblasts (HGFs) obtained from patients with healthy or inflammatory gingiva using DNA microarray analysis. We also studied the expression levels of these proteins by flow cytometric analysis (FACS). The expression levels of all eight genes in the HGFs of the Inflammatory group were significantly higher than those in the Healthy group on DNA microarray analysis. FACS revealed that the expression levels of all eight proteins on the HGFs of the Inflammatory group were higher than those on the Healthy group. Our data indicated that these eight proteins in HGFs are involved in inflammatory conditions in the gingiva, including periodontal disease. Our results suggested that these eight proteins, in turn, act directly or indirectly on the immune response by activating host cells involved in inflammatory processes. 相似文献