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Reiji Kannagi - One of the best experts on this subject based on the ideXlab platform.
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interaction of gata 3 t bet transcription factors regulates eXpression of sialyl lewis X homing receptors on th1 th2 lymphocytes
Proceedings of the National Academy of Sciences of the United States of America, 2006Co-Authors: Guoyun Chen, Hirotaka Osada, Luis F Santamariababi, Reiji KannagiAbstract:Selectin-dependent cell adhesion mediates inflammatory eXtravasation and routine homing of lymphocytes. Most resting peripheral T lymphocytes lack eXpression of sialyl Lewis X, the carbohydrate ligand for selectins, and are induced to strongly eXpress it upon activation. T helper 1 (Th1) cells are known to more preferentially eXpress sialyl Lewis X as compared with T helper 2 (Th2) cells upon activation. The molecular basis for this preferential eXpression, however, has not been elucidated to date. Here we show that the gene for fucosyltransferase VII (FUT7), the rate-limiting enzyme for sialyl Lewis X synthesis, is a unique eXample of the human genes with binding sites for both GATA-3 and T-bet, two opposing factors for Th1 and Th2 development, and is regulated transcriptionally by a balance of the two interacting transcription factors. T-bet promotes and GATA-3 represses FUT7 transcription. Our results indicated that T-bet interferes with the binding of GATA-3 to its target DNA, and also that GATA-3 significantly interferes with the binding of T-bet to the FUT7 promoter. T-bet has a binding ability to GATA-3, CBP/P300, and Sp1 to form a transcription factor compleX, and GATA-3 regulates FUT7 transcription by phosphorylation-dependently recruiting histone deacetylase (HDAC)-3/HDAC-5 and by competing with CBP/P300 in binding to the N terminus of T-bet. Suppression of GATA-3 activity by dominant-negative GATA-3 or repressor of GATA (ROG) was necessary to attain a maXimum eXpression of FUT7 and sialyl Lewis X in human T lymphoid cells. These results indicate that the GATA-3/T-bet transcription factor compleX regulates the cell-lineage-specific eXpression of the lymphocyte homing receptors.
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Interaction of GATA-3/T-bet transcription factors regulates eXpression of sialyl Lewis X homing receptors on Th1/Th2 lymphocytes.
Proceedings of the National Academy of Sciences of the United States of America, 2006Co-Authors: Guoyun Chen, Hirotaka Osada, Luis F. Santamaria-babi, Reiji KannagiAbstract:Selectin-dependent cell adhesion mediates inflammatory eXtravasation and routine homing of lymphocytes. Most resting peripheral T lymphocytes lack eXpression of sialyl Lewis X, the carbohydrate ligand for selectins, and are induced to strongly eXpress it upon activation. T helper 1 (Th1) cells are known to more preferentially eXpress sialyl Lewis X as compared with T helper 2 (Th2) cells upon activation. The molecular basis for this preferential eXpression, however, has not been elucidated to date. Here we show that the gene for fucosyltransferase VII (FUT7), the rate-limiting enzyme for sialyl Lewis X synthesis, is a unique eXample of the human genes with binding sites for both GATA-3 and T-bet, two opposing factors for Th1 and Th2 development, and is regulated transcriptionally by a balance of the two interacting transcription factors. T-bet promotes and GATA-3 represses FUT7 transcription. Our results indicated that T-bet interferes with the binding of GATA-3 to its target DNA, and also that GATA-3 significantly interferes with the binding of T-bet to the FUT7 promoter. T-bet has a binding ability to GATA-3, CBP/P300, and Sp1 to form a transcription factor compleX, and GATA-3 regulates FUT7 transcription by phosphorylation-dependently recruiting histone deacetylase (HDAC)-3/HDAC-5 and by competing with CBP/P300 in binding to the N terminus of T-bet. Suppression of GATA-3 activity by dominant-negative GATA-3 or repressor of GATA (ROG) was necessary to attain a maXimum eXpression of FUT7 and sialyl Lewis X in human T lymphoid cells. These results indicate that the GATA-3/T-bet transcription factor compleX regulates the cell-lineage-specific eXpression of the lymphocyte homing receptors.
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Design and synthesis of a novel neo-glycolipid containing sialyl Lewis X determinant carried on the mucin GlcNAcβ1-6GalNAcα core structure
Tetrahedron: Asymmetry, 2005Co-Authors: Nobumasa Otsubo, Reiji Kannagi, Hideharu Ishida, Makoto KisoAbstract:A novel neo-glycolipid containing sialyl Lewis X determinant carried on the mucin GlcNAcβ1-6GalNAcα core structure has been designed and synthesized. By employing this compound as a probe, the structure required for the recognition of anti-cancer antibodies, NCC-ST-439, has been elucidated.
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Molecular mechanism for cancer-associated induction of sialyl Lewis X and sialyl Lewis A eXpression-The Warburg effect revisited.
Glycoconjugate journal, 2004Co-Authors: Reiji KannagiAbstract:Cell adhesion mediated by selectins and their carbohydrate ligands, sialyl Lewis X and sialyl Lewis A, figures heavily in cancer metastasis. EXpression of these carbohydrate determinants is markedly enhanced in cancer cells, but the molecular mechanism that leads to cancer-associated eXpression of sialyl Lewis X/A has not been well understood. Results of recent studies indicated involvement of two principal mechanisms in the accelerated eXpression of sialyl Lewis X/A in cancers; ‘incomplete synthesis’ and ‘neosynthesis.’ As to ‘incomplete synthesis,’ we have recently found further modified forms of sialyl Lewis X and sialyl Lewis A in non-malignant colonic epithelium, which have additional 6-sulfation or 2 → 6 sialylation. The impairment of GlcNAc 6-sulfation and 2 → 6 sialylation upon malignant transformation leads to accumulation of sialyl Lewis X/A in colon cancer cells. Epigenetic changes such as DNA methylation and/or histone deacetylation are suggested to lie behind such incomplete synthesis. As to the mechanism called ‘neosynthesis,’ recent studies have indicated that cancer-associated alterations in the sugar transportation and intermediate carbohydrate metabolism play important roles. Cancer cells are known to eXhibit a metabolic shift from oXidative to elevated anaerobic glycolysis (Warburg effect), which is correlated with the increased gene eXpression of sugar transporters and glycolytic enzymes induced by common cancer-specific genetic alterations. The increased sialyl Lewis X/A eXpression in cancer is a link in the chains of these events because our recent results indicated that these events accompany transcriptional induction of a set of genes closely related to its eXpression. Published in 2004.
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Molecular mechanism for cancer-associated induction of sialyl Lewis X and sialyl Lewis A eXpression—The Warburg effect revisited
Glycoconjugate Journal, 2003Co-Authors: Reiji KannagiAbstract:Cell adhesion mediated by selectins and their carbohydrate ligands, sialyl Lewis X and sialyl Lewis A, figures heavily in cancer metastasis. EXpression of these carbohydrate determinants is markedly enhanced in cancer cells, but the molecular mechanism that leads to cancer-associated eXpression of sialyl Lewis X/A has not been well understood. Results of recent studies indicated involvement of two principal mechanisms in the accelerated eXpression of sialyl Lewis X/A in cancers; ‘incomplete synthesis’ and ‘ neo synthesis.’ As to ‘incomplete synthesis,’ we have recently found further modified forms of sialyl Lewis X and sialyl Lewis A in non-malignant colonic epithelium, which have additional 6-sulfation or 2 → 6 sialylation. The impairment of GlcNAc 6-sulfation and 2 → 6 sialylation upon malignant transformation leads to accumulation of sialyl Lewis X/A in colon cancer cells. Epigenetic changes such as DNA methylation and/or histone deacetylation are suggested to lie behind such incomplete synthesis. As to the mechanism called ‘ neo synthesis,’ recent studies have indicated that cancer-associated alterations in the sugar transportation and intermediate carbohydrate metabolism play important roles. Cancer cells are known to eXhibit a metabolic shift from oXidative to elevated anaerobic glycolysis (Warburg effect), which is correlated with the increased gene eXpression of sugar transporters and glycolytic enzymes induced by common cancer-specific genetic alterations. The increased sialyl Lewis X/A eXpression in cancer is a link in the chains of these events because our recent results indicated that these events accompany transcriptional induction of a set of genes closely related to its eXpression. Published in 2004.
Minoru Fukuda - One of the best experts on this subject based on the ideXlab platform.
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EXtended Core 1 and Core 2 Branched O-Glycans Differentially Modulate Sialyl Lewis X-type L-selectin Ligand Activity
The Journal of biological chemistry, 2003Co-Authors: Junya Mitoma, Jiunn-chern Yeh, John B Lowe, Bronislawa Petryniak, Nobuyoshi Hiraoka, Minoru FukudaAbstract:Abstract It has been established that sialyl Lewis X in core 2 branched O-glycans serves as an E- and P-selectin ligand. Recently, it was discovered that 6-sulfosialyl Lewis X in eXtended core 1 O-glycans, NeuNAcα2→3Galβ1→4(Fucα1→3(sulfo→6))GlcNAcβ1→ 3Galβ1→3GalNAcα1→Ser/Thr, functions as an L-selectin ligand in high endothelial venules. EXtended core 1O-glycans can be synthesized when a core 1 eXtension enzyme is present. In this study, we first show that β1,3-N-acetylglucosaminyltransferase-3 (β3GlcNAcT-3) is almost eXclusively responsible for core 1 eXtension among seven different β3GlcNAcTs and thus acts on core 1 O-glycans attached to PSGL-1. We found that transcripts encoding β3GlcNAcT-3 were eXpressed in human neutrophils and lymphocytes but that their levels were lower than those of transcripts encoding core 2 β1,6-N-acetylglucosaminyltransferase I (Core2GlcNAcT-I). Neutrophils also eXpressed transcripts encoding fucosyltransferase VII (FucT-VII) and Core2GlcNAcT-I, whereas lymphocytes eXpressed only small amounts of transcripts encoding FucT-VII. To determine the roles of sialyl Lewis X in eXtended core 1O-glycans, Chinese hamster ovary (CHO) cells were stably transfected to eXpress PSGL-1, FucT-VII, and either β3GlcNAcT-3 or Core2GlcNAcT-I. Glycan structural analyses disclosed that PSGL-1 eXpressed in these transfected cells carried comparable amounts of sialyl Lewis X in eXtended core 1 and core 2 branchedO-glycans. In a rolling assay, CHO cells eXpressing sialyl Lewis X in eXtended core 1 O-glycans supported a significant degree of shear-dependent tethering and rolling of neutrophils and lymphocytes, although less than CHO cells eXpressing sialyl Lewis X in core 2 branched O-glycans. These results indicate that sialyl Lewis X in eXtended core 1 O-glycans can function as an L-selectin ligand and is potentially involved in neutrophil adhesion on neutrophils bound to activated endothelial cells.
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Natural killer cells attack tumor cells eXpressing high levels of sialyl Lewis X oligosaccharides
Proceedings of the National Academy of Sciences of the United States of America, 2002Co-Authors: Chikara Ohyama, Michiko N Fukuda, Satoru Kanto, Kazunori Kato, Osamu Nakano, Yoichi Arai, Tetsuro Kato, Shihao Chen, Minoru FukudaAbstract:Epithelial carcinoma and leukemia cells eXpress sialyl Lewis X oligosaccharides as tumor-associated carbohydrate antigens. To determine the role of sialyl Lewis X oligosaccharides in tumor dissemination, human melanoma MeWo cells, which do not eXpress sialyl Lewis X, were transfected with α1,3-fucosyltransferase III (FTIII), and cell lines eXpressing different amounts of sialyl Lewis X were isolated. When these cells were injected into the tail vein of nude mice, cells eXpressing moderate amounts of sialyl Lewis X (MeWo-FTIII⋅M) produced a significantly greater number of lung tumor foci than did parental MeWo cells. In contrast, cells eXpressing large amounts of sialyl Lewis X (MeWo-FTIII⋅H) produced few lung tumor foci in nude mice but were highly tumorigenic in beige mice, which have defective natural killer (NK) cells. In vitro assays demonstrated that MeWo-FTIII⋅H cells are much more sensitive to NK cell-mediated cytotoXicity than are MeWo-FTIII⋅M cells or parental MeWo cells and the susceptibility of MeWo-FTIII⋅H cells to NK cell-mediated cytolysis can be inhibited by preincubating MeWo-FTIII⋅H cells with anti-sialyl Lewis X antibody. Moreover, we discovered that NK cell-mediated cytolysis of MeWo-FTIII⋅H cells can be inhibited by the addition of an antibody against the NK cell receptor CD94 or sialyl Lewis X oligosaccharides. These results, combined with structural analysis of MeWo-FTIII⋅H cell carbohydrates, indicate that moderate amounts of sialyl Lewis X lead to tumor metastasis, whereas eXpression of high levels of sialyl Lewis X leads to an NK cell attack on tumor cells, demonstrating that eXpression of different amounts of sialyl Lewis X results in entirely different biological consequences.
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sialyl lewis X dependent lung colonization of b16 melanoma cells through a selectin like endothelial receptor distinct from e or p selectin
Cancer Research, 2002Co-Authors: Chikara Ohyama, Minoru Fukuda, Jianing Zhang, Jun Nakayama, Masami Suzuki, Atsushi Suzuki, Michiko N FukudaAbstract:Endothelial carbohydrate binding proteins, E- and P-selectins, are thought to mediate sialyl Lewis A/X-dependent hematogenous cancer metastasis. We tested this hypothesis using sialyl Lewis X-dependent B16 melanoma lung targeting and its inhibition with selectin ligand mimicry peptide, IELLQAR. In E/P-selectin doubly deficient mutant mice, sialyl Lewis X-eXpressing B16 melanoma cells colonized the lung, and IELLQAR inhibited this colonization. However, tumors grown in E/P-selectin-deficient mice were significantly smaller than those grown in wild-type mice. These results indicate that the IELLQAR peptide receptor eXpressed in the lung vasculature plays a major role in sialyl Lewis X-dependent cancer cells targeting to the lung.
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a peptide mimic of e selectin ligand inhibits sialyl lewis X dependent lung colonization of tumor cells
Cancer Research, 2000Co-Authors: Michiko N Fukuda, Chikara Ohyama, Kevin P Lowitz, Osamu Matsuo, Renata Pasqualini, Erkki Ruoslahti, Minoru FukudaAbstract:Selectins bind to carbohydrate ligands in a calcium-dependent manner and play critical roles in host defense and possibly in tumor metastasis. To isolate peptides that mimic E-selectin ligands, we screened a phage peptide library using E-selectin as a target molecule. This attempt uneXpectedly failed, probably because the binding affinity of E-selectin to its ligand is low. We then took an approach that is analogous to the isolation of anti-idiotype antibodies and were able to isolate peptides that bound to anticarbohydrate antibodies recognizing E-selectin ligands. These peptides, enriched for their binding to anti-Lewis A antibody, were found to bind to E-, P- and L-selectins in a calcium-dependent manner. Phage harboring the identified peptide IELLQAR and synthetic peptides having the same sequence inhibited the binding of sialyl Lewis X or sialyl Lewis A oligosaccharides to E-selectin. The adhesion of HL-60 and B16 melanoma cells eXpressing sialyl Lewis X to E-selectin was also inhibited by the phage-displaying IELLQAR peptide. Moreover, i.v. injected IELLQAR peptide inhibited the lung colonization of mouse B16 melanoma and human lung tumor cells eXpressing sialyl Lewis X. These results demonstrate that it is possible to isolate peptides mimicking carbohydrate ligands by screening the peptides for binding to anticarbohydrate antibodies and then using them to inhibit carbohydrate-dependent eXperimental tumor metastasis.
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alpha(1,2)-fucosylation prevents sialyl Lewis X eXpression and E-selectin-mediated adhesion of fucosyltransferase VII-transfected cells.
European journal of biochemistry, 2000Co-Authors: Mourad Zerfaoui, Minoru Fukuda, Véronique Sbarra, Dominique Lombardo, Assou El-battariAbstract:E-selectin is a cytokine-inducible, calcium-dependent endothelial cell adhesion molecule that plays a critical role in the leucocyte-endothelium interaction during inflammation and is thought to contribute to the metastatic dissemination of tumour cells. Like the other selectins, E-selectin binds to ligands carrying the tetrasaccharide Sialyl-Lewis X (NeuAcalpha2,3Galbeta1,4[Fucalpha1, 3]GlcNAc)1 or its isomer Sialyl-Lewis a (NeuAcalpha2, 3Galbeta1, 3[Fucalpha1,4]GlcNAc). We eXamined the effect of eXpressing the H-type alpha(1,2)-fucosyltransferase or the alpha(2, 6)-sialyltransferase on the synthesis of Sialyl-Lewis X by alpha(1, 3)fucosyltransferase. We found that H-type alpha(1, 2)-fucosyltransferase but not alpha(2,6)-sialyltransferase, strongly inhibited Sialyl-Lewis X eXpression and E-selectin adhesion. We assume that H-type alpha(1,2)-fucosyltransferase competes with the endogenous alpha(2,3)-sialyltransferase for the N-acetyllactosamine structures assigned to further serve as acceptors for alpha(1, 3)fucosyltransferase.
Makoto Kiso - One of the best experts on this subject based on the ideXlab platform.
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Design and synthesis of a novel neo-glycolipid containing sialyl Lewis X determinant carried on the mucin GlcNAcβ1-6GalNAcα core structure
Tetrahedron: Asymmetry, 2005Co-Authors: Nobumasa Otsubo, Reiji Kannagi, Hideharu Ishida, Makoto KisoAbstract:A novel neo-glycolipid containing sialyl Lewis X determinant carried on the mucin GlcNAcβ1-6GalNAcα core structure has been designed and synthesized. By employing this compound as a probe, the structure required for the recognition of anti-cancer antibodies, NCC-ST-439, has been elucidated.
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specific detection of sialyl lewis X determinant carried on the mucin glcnacβ1 6galnacα core structure as a tumor associated antigen
Biochemical and Biophysical Research Communications, 1998Co-Authors: Kensuke Kumamoto, Makoto Kiso, Chikako Mitsuoka, Hideharu Ishida, Mineko Izawa, Naoko Kimura, Nobumasa Otsubo, Tesshi Yamada, Setsuo Hirohashi, Reiji KannagiAbstract:Abstract Sialyl Lewis X serves as a ligand for selectins and is proposed to be implicated in hematogenous metastasis of cancers. When a cultured human breast cancer cell line, MCF-7, which does not eXpress sialyl Lewis X, was transfected with human fucosyltransferase VI cDNA, a strong eXpression of sialyl Lewis X was induced on transfectant cells. The transfectant cells were found to be also reactive to the antibody NCC-ST-439, which was initially raised against human gastric cancer cells and later was shown to recognize a tumor-associated carbohydrate antigen in breast, gastric, and colon cancers. This suggested that the antigen recognized by NCC-ST-439 is closely related to sialyl Lewis X. Subsequent studies indicated that NCC-ST-439 specifically reacts to NeuAcα2→3Galβ1→4(Fucα1→3)GlcNAcβ1→6GalNAcα1→R, the sialyl Lewis X on the mucin GlcNAcβ1→6 GalNAcα structure. The antibody was not reactive to the conventional sialyl Lewis X determinants on straight and/or branched polylactosamine structures including NeuAcα2→3Galβ1→4(Fucα1→3)GlcNAcβ1→3Galβ1→4Glcβ1→R and NeuAcα2→3Galβ1→4(Fucα1→3)GlcNAcβ1→6Galβ1→4 Glcβ1→R. This was in clear contrast to most of the known anti-sialyl Lewis X antibodies, which do not discriminate internal structures carrying the sialyl Lewis X determinant. On the other hand, the newly generated monoclonal antibody GSC154-27 had a specificity completely the reverse of the specificity of NCC-ST-439 in that it was strongly reactive to the conventional sialyl Lewis X determinants in straight and branched polylactosamine structures, while far less reactive to the sialyl Lewis X determinant on the mucin GlcNAcβ1→6GalNAcα core structure. A set of these two antibodies would be useful in discriminating the molecular species of sialyl Lewis X eXpressed by malignant cells and in studying their functional significance.
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identification of a major carbohydrate capping group of the l selectin ligand on high endothelial venules in human lymph nodes as 6 sulfo sialyl lewis X
Journal of Biological Chemistry, 1998Co-Authors: Chikako Mitsuoka, Makoto Kiso, Hideharu Ishida, Mineko Izawa, Mikiko Sawadakasugai, Keiko Andofurui, Hayao Nakanishi, Shigeo Nakamura, Reiji KannagiAbstract:Abstract We investigated the molecular species of sulfated sialyl Lewis X determinants, the putative L-selectin ligand, eXpressed on high endothelial venules (HEV) in human lymph nodes. Comparison of the reactivity pattern of HEV with the reactivity of the pure 6-sulfo, 6′-sulfo, or 6,6′-bissulfo sialyl Lewis X determinant with hitherto known anti-sialyl Lewis X antibodies strongly suggested 6-sulfo sialyl Lewis X to be the best candidate for the major sulfated sialyl Lewis X determinant on HEV, followed by 6,6′-bissulfo sialyl Lewis X, whereas 6′-sulfo sialyl Lewis X was unlikely. We newly generated monoclonal antibodies (mAbs) G152 and G72 directed against 6-sulfo sialyl Lewis X, which intensely labeled HEV in immunohistochemical eXamination and inhibited binding of recombinant L-selectin-IgG to HEV, suggesting that the determinant serves as a ligand for L-selectin. To test the concomitant eXpression of 6,6′-bissulfo sialyl Lewis X, specific mAbs (G2706, G27011, G27037, and G27039) were generated, but all antibodies failed to react to HEV. NeXt, we established mAbs (AG97 and AG273) directed against 6-sulfo Lewis X, the asialo form of 6-sulfo sialyl Lewis X. The antibodies were not reactive to untreated HEV, but strongly reacted to sialidase-treated HEV. This indicated the predominance of the sialylated form of 6-sulfo sialyl Lewis X and minimal eXpression of its asialo form, corroborating that it was synthesized by fucosyltransferase VII, the isoenzyme that preferentially produces the sialylated form of the determinant.
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A Convenient and Efficient Synthesis of Sialyl Lewis X
Bioscience biotechnology and biochemistry, 1998Co-Authors: Kunihisa Baba, Hideharu Ishida, Akira Hasegawa, Noriyuki Iwata, Hitoshi Hamajima, Takao Ikami, Makoto KisoAbstract:A convenient synthesis of the sialyl Lewis X (sLeX) tetrasaccharide, NeuAcα2-3Galβ1-4(Fucα1-3)GlcNAc (8), as a carbohydrate ligand for selectins is described. The key step is the reaction between NeuAcα2-3GalSMe (5) as a glycosyl donor and the suitably protected Fucα1-3GlcNAc derivative (4) as the glycosyl acceptor by using dimethyl(methylthio)sulfonium triflate (DMTST) as the promoter.
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Sulfated Sialyl Lewis X, the Putative L-Selectin Ligand, Detected on Endothelial Cells of High Endothelial Venules by a Distinct Set of Anti-Sialyl Lewis X Antibodies
Biochemical and biophysical research communications, 1997Co-Authors: Chikako Mitsuoka, Makoto Kiso, Naoko Kawakami-kimura, Mikiko Kasugai-sawada, Nozomu Hiraiwa, Ken-ichi Toda, Hideharu Ishida, Akira Hasegawa, Reiji KannagiAbstract:Abstract Endothelial cells of high endothelial venules (HEV) in human peripheral lymph nodes eXpressed a distinct type of sialyl Lewis X antigen, which was detected preferentially with a set of anti-sialyl Lewis X antibodies, 2F3, 2H5 and HECA-452 in immunohistochemistry, while another set of anti-sialyl Lewis X antibodies, FH-6 and CSLEX-1, failed to detect it. The adhesion of cells eXpressing L-selectin to HEV was inhibited by members of the former set of antibodies in Stamper-Woodruff assays performed on frozen sections of human peripheral lymph nodes. Transfection of a cultured endothelial cell line with a human α1→3 fucosyltransferase, Fuc-T VII, resulted in the eXpression of a distinct type of sialyl Lewis X antigen having the reactivity similar to that of HEV;i.e.,the antigen appearing on the transfectant clone was detectable only with the set of 2F3, 2H5 and HECA-452, but not with the set of FH-6 and CSLEX-1. Treatment of transfectant cells with sodium chlorate, a metabolic inhibitor of sulfation, resulted in reactivity to the members of the latter set of antibodies, suggesting that sulfation of sialyl Lewis X moiety was the cause of the discrepancy in the reactivity of the anti-sialyl Lewis X antibodies. When tested against various authentic sulfated sialyl Lewis X determinants, 6-sulfo sialyl Lewis X and 6,6′-bis-sulfo sialyl Lewis X were found to be reactive to the antibodies, 2F3, 2H5 and HECA-452, but not with antibodies FH-6 and CSLEX-1, suggesting that the distinct type of sialyl Lewis X determinant on the HEV endothelial cells and Fuc-T VII-transfected endothelial cell clone are mainly 6-sulfo and/or 6,6′-bis-sulfo sialyl Lewis X determinants.
Hideharu Ishida - One of the best experts on this subject based on the ideXlab platform.
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Design and synthesis of a novel neo-glycolipid containing sialyl Lewis X determinant carried on the mucin GlcNAcβ1-6GalNAcα core structure
Tetrahedron: Asymmetry, 2005Co-Authors: Nobumasa Otsubo, Reiji Kannagi, Hideharu Ishida, Makoto KisoAbstract:A novel neo-glycolipid containing sialyl Lewis X determinant carried on the mucin GlcNAcβ1-6GalNAcα core structure has been designed and synthesized. By employing this compound as a probe, the structure required for the recognition of anti-cancer antibodies, NCC-ST-439, has been elucidated.
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specific detection of sialyl lewis X determinant carried on the mucin glcnacβ1 6galnacα core structure as a tumor associated antigen
Biochemical and Biophysical Research Communications, 1998Co-Authors: Kensuke Kumamoto, Makoto Kiso, Chikako Mitsuoka, Hideharu Ishida, Mineko Izawa, Naoko Kimura, Nobumasa Otsubo, Tesshi Yamada, Setsuo Hirohashi, Reiji KannagiAbstract:Abstract Sialyl Lewis X serves as a ligand for selectins and is proposed to be implicated in hematogenous metastasis of cancers. When a cultured human breast cancer cell line, MCF-7, which does not eXpress sialyl Lewis X, was transfected with human fucosyltransferase VI cDNA, a strong eXpression of sialyl Lewis X was induced on transfectant cells. The transfectant cells were found to be also reactive to the antibody NCC-ST-439, which was initially raised against human gastric cancer cells and later was shown to recognize a tumor-associated carbohydrate antigen in breast, gastric, and colon cancers. This suggested that the antigen recognized by NCC-ST-439 is closely related to sialyl Lewis X. Subsequent studies indicated that NCC-ST-439 specifically reacts to NeuAcα2→3Galβ1→4(Fucα1→3)GlcNAcβ1→6GalNAcα1→R, the sialyl Lewis X on the mucin GlcNAcβ1→6 GalNAcα structure. The antibody was not reactive to the conventional sialyl Lewis X determinants on straight and/or branched polylactosamine structures including NeuAcα2→3Galβ1→4(Fucα1→3)GlcNAcβ1→3Galβ1→4Glcβ1→R and NeuAcα2→3Galβ1→4(Fucα1→3)GlcNAcβ1→6Galβ1→4 Glcβ1→R. This was in clear contrast to most of the known anti-sialyl Lewis X antibodies, which do not discriminate internal structures carrying the sialyl Lewis X determinant. On the other hand, the newly generated monoclonal antibody GSC154-27 had a specificity completely the reverse of the specificity of NCC-ST-439 in that it was strongly reactive to the conventional sialyl Lewis X determinants in straight and branched polylactosamine structures, while far less reactive to the sialyl Lewis X determinant on the mucin GlcNAcβ1→6GalNAcα core structure. A set of these two antibodies would be useful in discriminating the molecular species of sialyl Lewis X eXpressed by malignant cells and in studying their functional significance.
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identification of a major carbohydrate capping group of the l selectin ligand on high endothelial venules in human lymph nodes as 6 sulfo sialyl lewis X
Journal of Biological Chemistry, 1998Co-Authors: Chikako Mitsuoka, Makoto Kiso, Hideharu Ishida, Mineko Izawa, Mikiko Sawadakasugai, Keiko Andofurui, Hayao Nakanishi, Shigeo Nakamura, Reiji KannagiAbstract:Abstract We investigated the molecular species of sulfated sialyl Lewis X determinants, the putative L-selectin ligand, eXpressed on high endothelial venules (HEV) in human lymph nodes. Comparison of the reactivity pattern of HEV with the reactivity of the pure 6-sulfo, 6′-sulfo, or 6,6′-bissulfo sialyl Lewis X determinant with hitherto known anti-sialyl Lewis X antibodies strongly suggested 6-sulfo sialyl Lewis X to be the best candidate for the major sulfated sialyl Lewis X determinant on HEV, followed by 6,6′-bissulfo sialyl Lewis X, whereas 6′-sulfo sialyl Lewis X was unlikely. We newly generated monoclonal antibodies (mAbs) G152 and G72 directed against 6-sulfo sialyl Lewis X, which intensely labeled HEV in immunohistochemical eXamination and inhibited binding of recombinant L-selectin-IgG to HEV, suggesting that the determinant serves as a ligand for L-selectin. To test the concomitant eXpression of 6,6′-bissulfo sialyl Lewis X, specific mAbs (G2706, G27011, G27037, and G27039) were generated, but all antibodies failed to react to HEV. NeXt, we established mAbs (AG97 and AG273) directed against 6-sulfo Lewis X, the asialo form of 6-sulfo sialyl Lewis X. The antibodies were not reactive to untreated HEV, but strongly reacted to sialidase-treated HEV. This indicated the predominance of the sialylated form of 6-sulfo sialyl Lewis X and minimal eXpression of its asialo form, corroborating that it was synthesized by fucosyltransferase VII, the isoenzyme that preferentially produces the sialylated form of the determinant.
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A Convenient and Efficient Synthesis of Sialyl Lewis X
Bioscience biotechnology and biochemistry, 1998Co-Authors: Kunihisa Baba, Hideharu Ishida, Akira Hasegawa, Noriyuki Iwata, Hitoshi Hamajima, Takao Ikami, Makoto KisoAbstract:A convenient synthesis of the sialyl Lewis X (sLeX) tetrasaccharide, NeuAcα2-3Galβ1-4(Fucα1-3)GlcNAc (8), as a carbohydrate ligand for selectins is described. The key step is the reaction between NeuAcα2-3GalSMe (5) as a glycosyl donor and the suitably protected Fucα1-3GlcNAc derivative (4) as the glycosyl acceptor by using dimethyl(methylthio)sulfonium triflate (DMTST) as the promoter.
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Sulfated Sialyl Lewis X, the Putative L-Selectin Ligand, Detected on Endothelial Cells of High Endothelial Venules by a Distinct Set of Anti-Sialyl Lewis X Antibodies
Biochemical and biophysical research communications, 1997Co-Authors: Chikako Mitsuoka, Makoto Kiso, Naoko Kawakami-kimura, Mikiko Kasugai-sawada, Nozomu Hiraiwa, Ken-ichi Toda, Hideharu Ishida, Akira Hasegawa, Reiji KannagiAbstract:Abstract Endothelial cells of high endothelial venules (HEV) in human peripheral lymph nodes eXpressed a distinct type of sialyl Lewis X antigen, which was detected preferentially with a set of anti-sialyl Lewis X antibodies, 2F3, 2H5 and HECA-452 in immunohistochemistry, while another set of anti-sialyl Lewis X antibodies, FH-6 and CSLEX-1, failed to detect it. The adhesion of cells eXpressing L-selectin to HEV was inhibited by members of the former set of antibodies in Stamper-Woodruff assays performed on frozen sections of human peripheral lymph nodes. Transfection of a cultured endothelial cell line with a human α1→3 fucosyltransferase, Fuc-T VII, resulted in the eXpression of a distinct type of sialyl Lewis X antigen having the reactivity similar to that of HEV;i.e.,the antigen appearing on the transfectant clone was detectable only with the set of 2F3, 2H5 and HECA-452, but not with the set of FH-6 and CSLEX-1. Treatment of transfectant cells with sodium chlorate, a metabolic inhibitor of sulfation, resulted in reactivity to the members of the latter set of antibodies, suggesting that sulfation of sialyl Lewis X moiety was the cause of the discrepancy in the reactivity of the anti-sialyl Lewis X antibodies. When tested against various authentic sulfated sialyl Lewis X determinants, 6-sulfo sialyl Lewis X and 6,6′-bis-sulfo sialyl Lewis X were found to be reactive to the antibodies, 2F3, 2H5 and HECA-452, but not with antibodies FH-6 and CSLEX-1, suggesting that the distinct type of sialyl Lewis X determinant on the HEV endothelial cells and Fuc-T VII-transfected endothelial cell clone are mainly 6-sulfo and/or 6,6′-bis-sulfo sialyl Lewis X determinants.
John B Lowe - One of the best experts on this subject based on the ideXlab platform.
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EXtended Core 1 and Core 2 Branched O-Glycans Differentially Modulate Sialyl Lewis X-type L-selectin Ligand Activity
The Journal of biological chemistry, 2003Co-Authors: Junya Mitoma, Jiunn-chern Yeh, John B Lowe, Bronislawa Petryniak, Nobuyoshi Hiraoka, Minoru FukudaAbstract:Abstract It has been established that sialyl Lewis X in core 2 branched O-glycans serves as an E- and P-selectin ligand. Recently, it was discovered that 6-sulfosialyl Lewis X in eXtended core 1 O-glycans, NeuNAcα2→3Galβ1→4(Fucα1→3(sulfo→6))GlcNAcβ1→ 3Galβ1→3GalNAcα1→Ser/Thr, functions as an L-selectin ligand in high endothelial venules. EXtended core 1O-glycans can be synthesized when a core 1 eXtension enzyme is present. In this study, we first show that β1,3-N-acetylglucosaminyltransferase-3 (β3GlcNAcT-3) is almost eXclusively responsible for core 1 eXtension among seven different β3GlcNAcTs and thus acts on core 1 O-glycans attached to PSGL-1. We found that transcripts encoding β3GlcNAcT-3 were eXpressed in human neutrophils and lymphocytes but that their levels were lower than those of transcripts encoding core 2 β1,6-N-acetylglucosaminyltransferase I (Core2GlcNAcT-I). Neutrophils also eXpressed transcripts encoding fucosyltransferase VII (FucT-VII) and Core2GlcNAcT-I, whereas lymphocytes eXpressed only small amounts of transcripts encoding FucT-VII. To determine the roles of sialyl Lewis X in eXtended core 1O-glycans, Chinese hamster ovary (CHO) cells were stably transfected to eXpress PSGL-1, FucT-VII, and either β3GlcNAcT-3 or Core2GlcNAcT-I. Glycan structural analyses disclosed that PSGL-1 eXpressed in these transfected cells carried comparable amounts of sialyl Lewis X in eXtended core 1 and core 2 branchedO-glycans. In a rolling assay, CHO cells eXpressing sialyl Lewis X in eXtended core 1 O-glycans supported a significant degree of shear-dependent tethering and rolling of neutrophils and lymphocytes, although less than CHO cells eXpressing sialyl Lewis X in core 2 branched O-glycans. These results indicate that sialyl Lewis X in eXtended core 1 O-glycans can function as an L-selectin ligand and is potentially involved in neutrophil adhesion on neutrophils bound to activated endothelial cells.
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molecular cloning of a cdna encoding a novel human leukocyte alpha 1 3 fucosyltransferase capable of synthesizing the sialyl lewis X determinant
Journal of Biological Chemistry, 1994Co-Authors: Shunji Natsuka, Kevin M Gersten, K Zenita, Reiji Kannagi, John B LoweAbstract:Abstract The sialyl Lewis X determinant (NeuAc alpha 2,3Gal beta 1, 4[Fuc alpha 1,3]GlcNAc) is an essential component of leukocyte counterreceptors for E-selectin and P-selectin. The final step in sialyl Lewis X synthesis is catalyzed by alpha-1,3-fucosyltransferases acting on sialylated glycoconjugate precursors. Cultured human leukocytic cell lines eXpress an alpha-1,3-fucosyltransferase gene termed Fuc-TIV or ELFT but do not eXpress the other three cloned human alpha-1,3-fucosyltransferase genes to any significant degree. The physiological role of Fuc-TIV/ELFT in sialyl Lewis X biosynthesis is uncertain, however, since it can catalyze the synthesis of this determinant in some, but not all, transfected cell lines in a manner that is dependent upon the glycosylation phenotype of the host cell. We report here the molecular cloning of a cDNA encoding a new human leukocyte alpha-1,3-fucosyltransferase, termed Fuc-TVII, capable of synthesizing the sialyl Lewis X moiety. The cDNA sequence predicts a 341-amino acid-long type II transmembrane protein typical of mammalian glycosyltransferases. When eXpressed in mammalian cells, the Fuc-TVII cDNA directs the synthesis of cell surface sialyl Lewis X moieties but not the Lewis X, Lewis a, sialyl Lewis a, or VIM-2 determinants. Fuc-TVII can efficiently utilize alpha-2,3-sialyllactosamine in vitro to form the sialyl Lewis X tetrasaccharide but does not utilize lactosamine to form the Lewis X moiety. Northern blot analyses show that the Fuc-TVII gene is transcribed in HL-60 cells, a human promyelocytic cell line, and in YT cells, a natural killer-like cell line. Fuc-TVII represents a leukocytic alpha-1,3-fucosyltransferase that can participate in selectin ligand synthesis via its ability to catalyze the synthesis of sialyl Lewis X determinants.
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molecular cloning of a fourth member of a human alpha 1 3 fucosyltransferase gene family multiple homologous sequences that determine eXpression of the lewis X sialyl lewis X and difucosyl sialyl lewis X epitopes
Journal of Biological Chemistry, 1992Co-Authors: Peter L Smith, John B Lowe, Robert KellyAbstract:We and others have previously described the isolation of three human alpha (1,3)fucosyltransferase genes which form the basis of a nascent glycosyltransferase gene family. We now report the molecular cloning and eXpression of a fourth homologous human alpha (1,3)fucosyltransferase gene. When transfected into mammalian cells, this fucosyltransferase gene is capable of directing eXpression of the Lewis X (Gal beta 1-->4[Fuc alpha 1-->3]GlcNAc), sialyl Lewis X (NeuNAc alpha 2-->3Gal beta 1-->4 [Fuc alpha 1-->3]GlcNAc), and difucosyl sialyl Lewis X (NeuNAc alpha 2-->3Gal beta 1-->4[Fuc alpha 1-->3]GlcNAc beta 1-->3 Gal beta 1-->4[Fuc alpha 1-->3]GlcNAc) epitopes. The enzyme shares 85% amino acid sequence identity with Fuc-TIII and 89% identity with Fuc-TV but differs substantially in its acceptor substrate requirements. Polymerase chain reaction analyses demonstrate that the gene is syntenic to Fuc-TIII and Fuc-TV on chromosome 19. Southern blot analyses of human genomic DNA demonstrate that these four alpha (1,3)fucosyltransferase genes account for all DNA sequences that cross-hybridize at low stringency with the Fuc-TIII catalytic domain. Using similar methods, a catalytic domain probe from Fuc-TIV identifies a new class of DNA fragments which do not cross-hybridize with the chromosome 19 fucosyltransferase probes. These results eXtend the molecular definition of a family of human alpha (1,3)fucosyltransferase genes and provide tools for eXamining fucosyltransferase gene eXpression.
