Molecular Cloning and Expression of cDNA Encoding Human 3′-Phosphoadenylylsulfate:galactosylceramide 3′-Sulfotransferase

Honke, Koichi; Tsuda, Masayuki; Hirahara, Yukie; Ishii, Atsushi; Makita, Akira; Wada, Yoshinao

doi:10.1074/jbc.272.8.4864

Cited by 114 publications

(100 citation statements)

References 25 publications

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“…2 and Fig. 3) has homology with the sequences found in other Golgi-associated sulfotransferases such as chick chondroitin sulfate 6-Osulfotransferase, human galactosylceramide sulfotransferase, hamster heparan sulfate 2-O-sulfotransferase, and human heparan sulfate 3-O-sulfotransferase (33)(34)(35)(36). In particular, the RDP sequence (residue 189 -191) is completely conserved, and hydrophobic amino acids are shared among these amino acid sequences (Fig.…”

Section: Isolation Of a Cdna Clone That Directs The Expression Of Thementioning

confidence: 87%

Expression Cloning of a Human Sulfotransferase That Directs the Synthesis of the HNK-1 Glycan on the Neural Cell Adhesion Molecule and Glycolipids

Ong¹,

Yeh

Ding

et al. 1998

Journal of Biological Chemistry

107

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The HNK-1 carbohydrate is expressed on various adhesion molecules in the nervous system and is suggested to play a role in cell-cell and cell-substratum interactions. Here we describe the isolation and functional expression of a cDNA encoding a human sulfotransferase that synthesizes the HNK-1 carbohydrate epitope. A mutant Chinese hamster ovary cell line, Lec2, which stably expresses human neural cell adhesion molecule (N-CAM) (Lec2-NCAM), was first established. Lec2-NCAM was co-transfected with a human fetal brain cDNA library, a cDNA encoding the rat glucuronyltransferase that forms a precursor of the HNK-1 carbohydrate, and a vector encoding the polyoma large T antigen. The transfected Lec2-NCAM cells expressing the HNK-1 glycan were enriched by fluorescence-activated cell sorting. Sibling selection of recovered plasmids resulted in a cDNA encoding a sulfotransferase, HNK-1ST, that directs the expression of the HNK-1 carbohydrate epitope on the cell surface. The deduced amino acid sequence indicates that the enzyme is a type II membrane protein. Sequence analysis revealed that there is a short amino acid sequence in the presumed catalytic domain, which is highly homologous to the corresponding sequence in other Golgi-associated sulfotransferases so far cloned. The amount of HNK-1ST transcript is high in fetal brain compared with fetal lung, kidney, and liver. Expression of HNK-1ST resulted in the formation of the HNK-1 epitope on N-CAM and a soluble chimeric form of HNK-1ST was shown to add a sulfate group to a precursor, GlcA␤133Gal␤134GlcNAc␤13 R, forming sulfo33GlcA␤133Gal␤134GlcNAc␤13 R. The results combined together indicate that the cloned HNK-1ST directs the synthesis of the HNK-1 carbohydrate epitope on both glycoproteins and glycolipids in the nervous tissues.

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Section: Isolation Of a Cdna Clone That Directs The Expression Of Thementioning

confidence: 87%

Expression Cloning of a Human Sulfotransferase That Directs the Synthesis of the HNK-1 Glycan on the Neural Cell Adhesion Molecule and Glycolipids

Ong¹,

Yeh

Ding

et al. 1998

Journal of Biological Chemistry

107

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“…Since the CGT-deficient mouse generates neither seminolipid nor its precursor, GalEAG, precise dissection of the functions of the precursor and its sulfated end product, seminolipid, is difficult. The gene of 3Ј-phosphoadenylylsulfate:galactosylceramide 3Ј-sulfotransferase (EC 2.8.2.11) that sulfates GalEAG to seminolipid, as well as GalCer to galactosylsulfatide, has recently been cloned (36,37). The anticipated sulfotransferase knockout mouse should be able to provide the definitive answer as to whether both GalEAG and seminolipid or only either GalEAG or seminolipid are essential for normal spermatogenesis.…”

Section: Discussionmentioning

confidence: 99%

Requirement of Seminolipid in Spermatogenesis Revealed by UDP-galactose:Ceramide Galactosyltransferase-deficient Mice

Fujimoto¹,

Tadano-Aritomi²,

Tokumasu³

et al. 2000

Journal of Biological Chemistry

101

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Although seminolipid has long been suspected to play an essential role in spermatogenesis because of its uniquely abundant and temporally regulated expression in the spermatocytes, direct experimental evidence has been lacking. We have tested the hypothesis by examining the testis of the UDP-galactose:ceramide galactosyltransferase-deficient mouse, which is incapable of synthesizing seminolipid. Spermatogenesis in homozygous affected males is arrested at the late pachytene stage and the spermatogenic cells degenerate through the apoptotic process. This stage closely follows the phase of rapid seminolipid synthesis in the wild-type mouse. These observations not only provide the first experimental evidence that seminolipid is indeed essential for normal spermatogenesis but also support the broader concept that cell surface glycolipids are important in cellular differentiation and cell-to-cell interaction.Seminolipid (3-sulfogalactosyl-1-alkyl-2-acyl-sn-glycerol) is the principal glycolipid in spermatozoa of mammals comprising, for example, approximately 3% of total lipids and more than 90% of total glycolipids in boar spermatozoa (1-3). During spermatogenesis, seminolipid is synthesized rapidly in the early phase of spermatocyte development and maintained in subsequent germ cell stages (4 -6). This developmentally regulated rapid synthesis suggested a specific and possibly essential function of seminolipid in spermatogenesis (7) but experimental evidence has been lacking. Firm evidence in support of the speculation would have important bearing to the general concept that cell surface glycoconjugates are important in cellular differentiation, and cell-to-cell interaction (8).Seminolipid is synthesized by sulfation of its precursor, galactosylalkylacylglycerol (GalEAG) 1 . GalEAG is synthesized by UDPgalactose:ceramide galactosyltransferase (CGT, EC 2.4.1.62), which, besides GalEAG, also synthesizes the major myelin galactolipid, galactosylceramide (GalCer), galactosylsphingosine (psychosine), and galactosyldiacylglycerol (GalAAG) (9, 10). The CGTdeficient mice recently generated by gene-targeting do not synthesize any of these products and subsequent derivatives of the products (11)(12)(13)(14). Thus, the CGT-deficient mouse is an ideal experimental model to examine the consequences of lack of seminolipid to spermatogenesis. This report describes the first definitive evidence that deficient seminolipid biosynthesis indeed causes devastating disruption of the normal spermatogenetic process. EXPERIMENTAL PROCEDURESMice-The mice heterozygous for the disrupted Cgt gene (11) were originally supplied by Dr. B. Popko and maintained by backcrossing to C57BL/6N. Genotype was determined according to Coetzee et al. (11). WBB6F1 Kit W/W-v and WBB6F1 Mg f Sl/Sl-d mutant mice were purchased from Japan SLC, Inc., and C57BL/6N inbred mice were purchased from CLEA Japan, Inc. Isolation of Testicular Germ Cells-Testicular germ cells were isolated from decapsulated testes of sexually mature male C57BL/6N mice (15).RT-PCR Analysis-RNA...

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“…13) cDNA cloning of Gal3ST-1 The cDNA encoding for Gal3ST-1 9) was amplified from "Super Script" human testis cDNA library (Life Technologies, Inc., Rockville, MD) by PCR. Oligonucleotide primers used for the PCR were 5′-tttaagcttGTCTGAGATGCTGCCA-3′ (forward primer) and 5′-tttggatccTGGGACGTCACCACCG-3′ (reverse primer).…”

Section: )mentioning

confidence: 99%

“…Four Gal3ST genes have so far been identified [9][10][11][12][13] and their genetic and biochemical characters are summarized in Table I. Honke et al 9,14) purified cerebroside 3′-sulfotransferase (Gal3ST-1) from human renal cancer cells and isolated its cDNA based on partial amino acid sequences. Gal3ST-2 has broad substrate specificity and is expressed in various tissues including colonic epithelial cells.…”

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confidence: 99%

Down‐regulation of Gal 3–O‐sulfotransferase‐2 (Gal3ST‐2) Expression in Human Colonic Non‐mucinous Adenocarcinoma

Seko¹,

Nagata²,

Yonezawa³

et al. 2002

Japanese Journal of Cancer Research

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Expression levels of sulfomucin in human colonic adenocarcinomas are lower than those in normal colonic mucosa; this should be in part caused by down-regulation of expression of sulfotransferases, but it remains unclear which Gal 3-O-sulfotransferase (Gal3ST) is responsible for the biosynthesis of sulfomucin. In this study, we first examined the substrate specificities of four Gal3STs cloned so far, and found that Galβ β β β1 → → → →3GlcNAcβ β β β1 → → → → 3Galβ β β β1 → → → →4Glc (LNT) can be utilized only byGal3ST-2 as an acceptor substrate. The substrate specificity of Gal3ST-2 is closely similar to those of Gal3ST activities present in human normal mucosa and adenocarcinomas, suggesting that Gal3ST-2 is the dominant Gal3ST in colon and colonic cancer. Secondly, using LNT as a substrate, we comparatively analyzed levels of Gal3ST-2 activities in non-mucinous adenocarcinoma, mucinous adenocarcinomas, and the adjacent normal mucosa. We found that levels of Gal3ST-2 activities in non-mucinous adenocarcinoma are significantly lower than those in the adjacent normal mucosa, while those in mucinous adenocarcinomas are not significantly different from those in the adjacent normal mucosa. Moreover, we showed by a competitive RT-PCR method that expression levels of transcript for Gal3ST-2 in non-mucinous adenocarcinoma are lower than those in normal mucosa. These results suggest that Gal3ST-2 is one of the enzymes responsible for biosynthesis of sulfomucin, and that expression levels of Gal3ST-2 are down-regulated in non-mucinous adenocarcinoma.Key words: Colon cancer -Sulfomucin -Sulfotransferase -Gal3ST -Type 1Sulfomucin is a glycoprotein containing large amounts of O-linked, sulfated glycans. It is well-known that expression levels of sulfomucin in colonic cancer are markedly lower than those in the adjacent normal mucosa. 1, 2)Yamori et al.3) explored a monoclonal antibody, 91.9H, recognizing sulfated glycans in sulfomucin, and Irimura et al. 4) and Matsushita et al. 5) showed that expression levels of the epitope in colonic adenocarcinomas are indeed lower than those in normal mucosa. Thereafter, Loveless et al. 6) showed that the minimum epitope moiety is the SO 3 − →3Galβ1→3(Fucα1→4)GlcNAcβ1→3Galβ1→ structure. These results suggest that the decrease of the epitope in colon cancer is at least partly associated with that of sulfomucin. We previously showed that levels of the enzymatic activities of β1,3-galactosyltransferase, which can synthesize the backbone structure of the 91.9H antigen, are lower in colonic adenocarcinomas than in the adjacent normal mucosa. 7) Thereafter, Salvini et al. 8) showed that expression of β3GalT-V gene is down-regulated in colonic adenocarcinomas, suggesting that β3GalT-V gene is involved in biosynthesis of the 91.9H antigen and its low expression in colonic adenocarcinomas. However, it remains unclear whether or not 3-O-sulfation at the Gal residue is also down-regulated in the course of colonic carcinogenesis. Gal3ST transfers sulfate from PAPS to the C-3 position of Gal residu...

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Molecular Cloning and Expression of cDNA Encoding Human 3′-Phosphoadenylylsulfate:galactosylceramide 3′-Sulfotransferase

Cited by 114 publications

References 25 publications

Expression Cloning of a Human Sulfotransferase That Directs the Synthesis of the HNK-1 Glycan on the Neural Cell Adhesion Molecule and Glycolipids

Expression Cloning of a Human Sulfotransferase That Directs the Synthesis of the HNK-1 Glycan on the Neural Cell Adhesion Molecule and Glycolipids

Requirement of Seminolipid in Spermatogenesis Revealed by UDP-galactose:Ceramide Galactosyltransferase-deficient Mice

Down‐regulation of Gal 3–O‐sulfotransferase‐2 (Gal3ST‐2) Expression in Human Colonic Non‐mucinous Adenocarcinoma

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