Purification and Characterization of UDP-N-Acetylglucosamine: α1,3-d-Mannoside β1,4-N-Acetylglucosaminyltransferase (N-Acetylglucosaminyltransferase-IV) from Bovine Small Intestine

Oguri, Suguru; Minowa, Mari T.; Ihara, Yoshito; Taniguchi, Naoyuki; Ikenaga, Hiroshi; Takeuchi, Makoto

doi:10.1074/jbc.272.36.22721

Cited by 65 publications

(51 citation statements)

References 34 publications

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“…4C). As shown in earlier substrate specificity studies (2,3) or kinetic analysis (4), it has been reported that GnT-IV and GnT-V are unable to act on bisected sugar chains in vitro, whereas GnT-III is capable of reacting with any types of chain, i.e. bi-, tri-, or tetra-antennary.…”

Section: Resultsmentioning

confidence: 89%

“…Cell Culture-Huh6 cells were grown and maintained at 37°C in Dulbecco's modified Eagle's medium containing 4.5 g/liter of glucose (Nikken) supplemented with 10% fetal calf serum (Invitrogen), 50 units/ml penicillin G, and 50 g/ml streptomycin under a humidified atmosphere of 95% air and 5% CO 2 .…”

Section: Methodsmentioning

confidence: 99%

“…Oligosaccharides containing a bisecting GlcNAc (bisected oligosaccharides) do not serve as substrates for GnT-IV and GnT-V, which catalyze the formation of a ␤1,4-branch at an ␣1,3Man and a ␤1,6-branch at an ␣1,6Man, respectively (2,3). We previously reported that the action of GnT-V was strictly inhibited by the addition of a bisecting GlcNAc at the catalytic step (4).…”

Section: N-acetylglucosaminyltransferase III (Gnt-iii)mentioning

confidence: 99%

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Caveolin-1 Regulates the Functional Localization of N-Acetylglucosaminyltransferase III within the Golgi Apparatus

Sasai¹,

Ikeda

Ihara

et al. 2003

Journal of Biological Chemistry

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In an investigation of the mechanism underlying the functional sublocalization of glycosyltransferases within the Golgi apparatus, caveolin-1 was identified as a possible cellular factor. Caveolin-1 appears to regulate the localization of N-acetylglucosaminyltransferase III (GnT-III) in the intra-Golgi subcompartment. Structural analyses of total cellular N-glycans indicated that the overexpression of GnT-III in human hepatoma cells, in which caveolin-1 is not expressed, failed to reduce branch formation, whereas expression of caveolin-1 led to a dramatic decrease in the extent of branching with no enhancement in GnT-III activity. Because the addition of a bisecting GlcNAc by GnT-III to the core ␤-Man in N-glycans prevents the action of GnT-IV and GnT-V, both of which are involved in branch formation, this result suggests that caveolin-1 facilitates the prior action of GnT-III, relative to the other GnTs, on the nascent sugar chains in the Golgi apparatus and that GnT-III is redistributed in the earlier Golgi subcompartment by caveolin-1. Indeed, when caveolin-1 was expressed in human hepatoma cells, it was found to be co-localized with GnT-III, as evidenced by the fractionation of Triton X-100-insoluble cellular membranes by density gradient ultracentrifugation. Caveolin-1 may modify the biosynthetic pathway of sugar chains via the regulation of the intra-Golgi subcompartment localization of this key glycosyltransferase. N-Acetylglucosaminyltransferase III (GnT-III)1 is one of the Golgi-resident glycosyltransferases that catalyzes the transfer of GlcNAc from an UDP-GlcNAc to a core ␤-mannosyl residue of asparagine-linked glycans (N-glycans) via a ␤1,4-linkage, resulting in the formation of a unique structure, a bisecting GlcNAc (1). Oligosaccharides containing a bisecting GlcNAc (bisected oligosaccharides) do not serve as substrates for GnT-IV and GnT-V, which catalyze the formation of a ␤1,4-branch at an ␣1,3Man and a ␤1,6-branch at an ␣1,6Man, respectively (2, 3). We previously reported that the action of GnT-V was strictly inhibited by the addition of a bisecting GlcNAc at the catalytic step (4). Therefore, the action of GnT-III prior to these enzymes in the Golgi leads to an inhibition of the biosynthesis of multiantennary oligosaccharides. Indeed, the ectopic expression of GnT-III caused an effective reduction in multiantennary oligosaccharides in various types of cells (5, 6). For example, in the case of swine endothelial cells, the expression of GnT-III led to a dramatic decrease in multiantennary oligosaccharides, accompanied by a marked increase in bisected biantennary ones (6). However, there are exceptions, in which a high GnT-III activity did not lead to a decrease in branching. In the case of human hepatoma cells, the overexpression of GnT-III resulted in a dramatic increase in bisected oligosaccharides but did not cause a decrease of the number of antenna (7). It would be expected that GnT-IV and GnT-V act prior to GnT-III in Huh6 and HepG2 cells, because the activities of GnT-III, GnT-IV, and Gn...

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Section: Resultsmentioning

confidence: 89%

Section: Methodsmentioning

confidence: 99%

Section: N-acetylglucosaminyltransferase III (Gnt-iii)mentioning

confidence: 99%

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Caveolin-1 Regulates the Functional Localization of N-Acetylglucosaminyltransferase III within the Golgi Apparatus

Sasai¹,

Ikeda

Ihara

et al. 2003

Journal of Biological Chemistry

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“…N-glycan modifications mediated by GnTs greatly influence the invasion and migration abilities of cancer cells (6,7). N-acetylglucosaminyltransferase IV (GnT-IV) catalyses the formation of β1,4GlcNAc branches on the mannose core of N-glycans (8,9). Many previous studies showed that GnT is involved in the progression of cancers of the liver, pancreas, kidney and colon (10)(11)(12)(13).…”

Section: Introductionmentioning

confidence: 99%

N-acetylglucosaminyltransferase IVa promotes invasion of choriocarcinoma

et al. 2017

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Abstract. Gestational trophoblastic neoplasia (GTN)results from the malignant transformation of placental trophoblasts which secrete human chorionic gonadotropin (hCG) as do normal placenta or hydatidiform mole. N-acetylglucosaminyltransferase IV (GnT-IV) is a glycosyltransferase which catalyses the formation of β1,4GlcNAc branches on the mannose core of N-glycans. Previous studies reported that β1,4GlcNAc branches on hCG were detected in GTN but not in normal pregnancy or hydatidiform mole. The aim of the present study was to understand the role of GnT-IVa in choriocarcinoma and find the target proteins for GnT-IVa glycosylation which contribute to the malignancy of choriocarcinoma. Immunohistochemistry showed that Griffonia simplicifolia lectin-II staining and GnT-IVa staining were intense in trophoblastic cells of invasive mole and choriocarcinoma. We established a choriocarcinoma cell line with GnT-IVa overexpression (Jar-GnT4a), and examined its malignant potential and target proteins for GnT-IVa glycosylation. GnT-IVa overexpression increased the cell migration and invasion (2.5-and 1.4-fold) as well as the ability to adhere to the extracellular matrix (ECM) components, including fibronectin and collagen type I and IV. The tumour formation potential of Jar-GnT4a in mice was significantly higher than that of control (P= 0.0407), and the cumulative survival rate of mice with Jar-GnT4a was relatively lower than those with control. Immunoprecipitation studies showed that β1,4GlcNAc branches of N-glycans on integrin β1 in choriocarcinoma cells were increased by GnT-IVa overexpression. Nano-LC/MS/MS analysis suggested that lysosome-associated membrane glycoprotein 2 (LAMP-2) was a target protein for glycosylation by GnT-IVa. The increase in β1,4GlcNAc branches on LAMP-2 by GnT-IVa overexpression was confirmed by lectin blot analysis using whole cell lysate and conditioned medium. Our results suggest that highly branched N-glycans generated by the action of GnT-IVa are present in trophoblastic cells of GTN in proportion to GnT-IVa expression level, and that GnT-IVa may contribute to the malignancy of choriocarcinoma by promoting cell adhesion, migration and invasion through glycosylation of integrin β1 and LAMP-2.

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“…18) Subsequently, this strategy was applied to the other enzymes such as GnT-V 20),21) GnT-IV, 22) GnT-VI, 23),24) the central type I synthetic enzyme, 25) and α1-6 fucosyltransferase (Fut8) 26),27) (Fig. 1).…”

mentioning

confidence: 99%

Functional glycomics and evidence for gain- and loss-of-functions of target proteins for glycosyltransferases involved in <i>N</i>-glycan biosynthesis: their pivotal roles in growth and development, cancer metastasis and antibody therapy against cancer

Taniguchi

Takahashi

et al. 2004

Proceedings of the Japan Academy. Ser. B: Physical and Biologic

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N -Acetylglucosaminyltransferase V (GnT-V), N -acetylglucosaminyltransferase III (GnT-III) and α 1-6 fucosyltransferase (Fut8) catalyze reactions that form biologically important branching N -linked sugar chains in glycoproteins. The above three branching N -glycan sugar chains, β 1-6 GlcNAc branching, bisecting GlcNAc and core fucose ( α 1-6 fucose), play major roles in cancer invasion and metastasis, the inhibition of cancer metastasis, and antibody-dependent cellular cytotoxicity (ADCC), growth and development, respectively. A functional glycomic approach identified the gain- and loss-of-functions of glycoproteins as the result of the aberrant glycosylation. A membrane-type metal dependent serine proteinase designated matriptase which contains β 1-6 GlcNAc branching became resistant to auto-digestion and proteolysis by trypsin, and resulted in a constitutively active form which might be implicated in cancer invasion and metastasis. GnT-V also acts as an angiogenic factor without the mediation of functions as a glycosyltransferase. Recently, a GnT-V homologue, GnT-IX has been identified. This gene is expressed at relatively high levels in the brain and acts on N -glycans to form a unique branched structure, as well as O -mannosyl glycans. The addition of bisecting GlcNAc to various signaling molecules or adhesion molecules suppresses cancer metastasis. Fut8 knock-out mice, due to the lack of a core fucose ( α 1-6 fucose) in target glycoproteins, show disorders in growth and development. The presence of a bisecting GlcNAc or the absence of a core fucose in IgG molecules enhances ADCC activity for killing tumor cells by up to 10 to 100 fold and therefore is thought to have considerable use in antibody therapy against cancer. These data clearly indicate that gain- and loss-of-functions of target proteins for these glycosyltransferases are biologically important.

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Purification and Characterization of UDP-N-Acetylglucosamine: α1,3-d-Mannoside β1,4-N-Acetylglucosaminyltransferase (N-Acetylglucosaminyltransferase-IV) from Bovine Small Intestine

Cited by 65 publications

References 34 publications

Caveolin-1 Regulates the Functional Localization of N-Acetylglucosaminyltransferase III within the Golgi Apparatus

Caveolin-1 Regulates the Functional Localization of N-Acetylglucosaminyltransferase III within the Golgi Apparatus

N-acetylglucosaminyltransferase IVa promotes invasion of choriocarcinoma

Functional glycomics and evidence for gain- and loss-of-functions of target proteins for glycosyltransferases involved in <i>N</i>-glycan biosynthesis: their pivotal roles in growth and development, cancer metastasis and antibody therapy against cancer

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