Publications

• Dermatan 4-O-sulfotransferase 1 is pivotal in the formation of iduronic acid blocks in dermatan sulfate.

  Pacheco B., Maccarana M., Malmstrom A.

  Chondroitin/dermatan sulfate is a highly complex linear polysaccharide ubiquitously found in the extracellular matrix and at the cell surface. Several of its functions, such as binding to growth factors, are mediated by domains composed of alternating iduronic acid and 4-O-sulfated N-acetylgalacto ... >> More

  Chondroitin/dermatan sulfate is a highly complex linear polysaccharide ubiquitously found in the extracellular matrix and at the cell surface. Several of its functions, such as binding to growth factors, are mediated by domains composed of alternating iduronic acid and 4-O-sulfated N-acetylgalactosamine residues, named 4-O-sulfated iduronic acid blocks. These domains are generated by the action of two DS-epimerases, which convert D-glucuronic acid into its epimer L-iduronic acid, in close connection with 4-O-sulfation. In this study, dermatan sulfate structure was evaluated after downregulating or increasing dermatan 4-O-sulfotransferase 1 (D4ST-1) expression. siRNA-mediated downregulation of D4ST-1 in primary human lung fibroblasts led to a drastic specific reduction of iduronic acid blocks. No change of epimerase activity was found, indicating that the influence of D4ST-1 on epimerization is not due to an altered expression level of the DS-epimerases. Analysis of the dermatan sulfate chains showed that D4ST-1 is essential for the biosynthesis of the disulfated structure iduronic acid-2-O-sulfate-N-acetylgalactosamine-4-O-sulfate, thus confirmed to be strictly connected with the iduronic acid blocks. Also the biologically important residue hexuronic acid-N-acetylgalactosamine-4,6-O-disulfate considerably decreased after D4ST-1 downregulation. In conclusion, D4ST-1 is a key enzyme and is indispensable in the formation of important functional domains in dermatan sulfate and cannot be compensated by other 4-O-sulfotransferases. << Less

  Glycobiology 19:1197-1203(2009) [PubMed] [EuropePMC]

• Specificities of three distinct human chondroitin/dermatan N-acetylgalactosamine 4-O-sulfotransferases demonstrated using partially desulfated dermatan sulfate as an acceptor. Implication of differential roles in dermatan sulfate biosynthesis.

  Mikami T., Mizumoto S., Kago N., Kitagawa H., Sugahara K.

  4-O-Sulfation of GalNAc is a high frequency modification of chondroitin sulfate and dermatan sulfate (DS), and three major GalNAc 4-O-sulfotransferases including dermatan 4-O-sulfotransferase-1 (D4ST-1) and chondroitin 4-O-sulfotransferases-1 and -2 (C4ST-1 and -2) have been identified. 4-O-Sulfat ... >> More

  4-O-Sulfation of GalNAc is a high frequency modification of chondroitin sulfate and dermatan sulfate (DS), and three major GalNAc 4-O-sulfotransferases including dermatan 4-O-sulfotransferase-1 (D4ST-1) and chondroitin 4-O-sulfotransferases-1 and -2 (C4ST-1 and -2) have been identified. 4-O-Sulfation of GalNAc during DS biosynthesis had long been postulated to be a prerequisite for iduronic acid (IdoUA) formation by C5-epimerization of GlcUA. This hypothesis has recently been argued based on enzymological studies using microsomes that C5-epimerization precedes 4-O-sulfation, which was further supported by the specificity of the cloned D4ST-1 with predominant preference for IdoUA-GalNAc flanked by GlcUA-GalNAc over IdoUA-GalNAc flanked by IdoUA-GalNAc in exhaustively desulfated dermatan. Whereas the counterproposal explains the initial reactions, apparently it cannot rationalize the synthetic mechanism of IdoUA-GalNAc(4-O-sulfate)-rich clusters typical of mature DS chains. In this study, we examined detailed specificities of the three recombinant human 4-O-sulfotransferases using partially desulfated DS as an acceptor. Enzymatic analysis of the transferase reaction products showed that D4ST-1 far more efficiently transferred sulfate to GalNAc residues in -IdoUA-Gal-NAc-IdoUA-than in -GlcUA-GalNAc-GlcUA-sequences. In contrast, C4ST-1 showed the opposite preference, and C4ST-2 used GalNAc residues in both sequences to comparable degrees, being consistent with its phylogenetic relations to D4ST-1 and C4ST-1. Structural analysis of the oligosaccharides, which were isolated after chondroitinase AC-I digestion of the 35S-labeled transferase reaction products, revealed for the first time that D4ST-1, as compared with C4ST-1 and C4ST-2, most efficiently utilized GalNAc residues located not only in the sequence -IdoUA-GalNAc-IdoUA- but also in -GlcUA-Gal-NAc-IdoUA- and -IdoUA-GalNAc-GlcUA-. The isolated oligosaccharide structures also suggest that 4-O-sulfation promotes subsequent 4-O-sulfation of GalNAc in the neighboring disaccharide unit. << Less

  J. Biol. Chem. 278:36115-36127(2003) [PubMed] [EuropePMC]

• Molecular cloning and characterization of a dermatan-specific N-acetylgalactosamine 4-O-sulfotransferase.

  Evers M.R., Xia G., Kang H.-G., Schachner M., Baenziger J.U.

  We have identified and characterized an N-acetylgalactosamine-4-O-sulfotransferase designated dermatan-4-sulfotransferase-1 (D4ST-1) (GenBank(TM) accession number AF401222) based on its homology to HNK-1 sulfotransferase. The cDNA predicts an open reading frame encoding a type II membrane protein ... >> More

  We have identified and characterized an N-acetylgalactosamine-4-O-sulfotransferase designated dermatan-4-sulfotransferase-1 (D4ST-1) (GenBank(TM) accession number AF401222) based on its homology to HNK-1 sulfotransferase. The cDNA predicts an open reading frame encoding a type II membrane protein of 376 amino acids with a 43-amino acid cytoplasmic domain and a 316-amino acid luminal domain containing two potential N-linked glycosylation sites. D4ST-1 has significant amino acid identity with HNK-1 sulfotransferase (21.4%), N-acetylgalactosamine-4-O-sulfotransferase 1 (GalNAc-4-ST1) (24.7%), N-acetylgalactosamine-4-O-sulfotransferase 2 (GalNAc-4-ST2) (21.0%), chondroitin-4-O-sulfotransferase 1 (27.3%), and chondroitin-4-O-sulfotransferase 2 (22.8%). D4ST-1 transfers sulfate to the C-4 hydroxyl of beta1,4-linked GalNAc that is substituted with an alpha-linked iduronic acid (IdoUA) at the C-3 hydroxyl. D4ST-1 shows a strong preference in vitro for sulfate transfer to IdoUAalpha1,3GalNAcbeta1,4 that is flanked by GlcUAbeta1,3GalNAcbeta1,4 as compared with IdoUAalpha1,3GalNAcbeta1,4 flanked by IdoUAalpha1,3GalNAcbeta1,4. The specificity of D4ST-1 when assayed in vitro suggests that the addition of sulfate to GalNAc occurs immediately after epimerization of GlcUA to IdoUA. The open reading frame of D4ST-1 is encoded by a single exon located on human chromosome 15q14. Northern blot analysis reveals a single 2.4-kilobase transcript. D4ST-1 message is expressed in virtually all tissues at some level but is most highly expressed in pituitary, placenta, uterus, and thyroid. The properties of D4ST-1 indicate that sulfation of the GalNAc moieties in dermatan is mediated by a distinct GalNAc-4-O-sulfotransferase and occurs following epimerization of GlcUA to IdoUA. << Less

  J. Biol. Chem. 276:36344-36353(2001) [PubMed] [EuropePMC]

• Chondroitin sulfate/dermatan sulfate hybrid chains in the development of cerebellum. Spatiotemporal regulation of the expression of critical disulfated disaccharides by specific sulfotransferases.

  Mitsunaga C., Mikami T., Mizumoto S., Fukuda J., Sugahara K.

  Chondroitin sulfate/dermatan sulfate (CS/DS) chains regulate the development of the central nervous system in vertebrates. Previously, we demonstrated that CS/DS hybrid chains from embryonic pig brain exhibit neuritogenic and growth factor binding activities, which depended on their IdoUA content ... >> More

  Chondroitin sulfate/dermatan sulfate (CS/DS) chains regulate the development of the central nervous system in vertebrates. Previously, we demonstrated that CS/DS hybrid chains from embryonic pig brain exhibit neuritogenic and growth factor binding activities, which depended on their IdoUA content defining the DS-like structure. To elucidate the distribution of such functional sugar chains during the development of the brain, in situ hybridization was performed to examine expression of three CS/DS GalNAc 4-O-sulfotransferases, D4ST-1, C4ST-1, and C4ST-2, and a single uronyl 2-O-sulfotransferase (UST) involved in the biosynthesis of DS in addition to CS intermediates. C4ST-1 and C4ST-2 were ubiquitously expressed in the postnatal mouse brain, whereas the expression of D4ST-1 and UST was restricted in the developing cerebellum and culminated at postnatal day 14 as shown by reverse transcriptase-PCR analysis. In situ analysis of the disaccharides of CS/DS in brain sections revealed that the concentration of CS/DS increases 2-fold during development (postnatal day 7 to 7 weeks). The proportions of DS-specific, principal disaccharides, IdoUA-Gal-NAc(4-O-sulfate) (iA) and IdoUA(2-O-sulfate)-GalNAc(4-O-sulfate) (iB), produced by the sequential actions of D4ST-1 and UST, were higher in the CS/DS chains from cerebellum than those from whole brain sections. A dramatic increase (10-fold) in the proportion of iB during development was noteworthy. In contrast, GlcUA/IdoUA(2-O-sulfate)-GalNAc(6-O-sulfate) (D/iD) and GlcUA/IdoUA-GalNAc(4, 6-O-disulfate) (E/iE) decreased to 50 and 30%, respectively, in the developing cerebellum. These results suggest that the IdoUA-containing iA and iB units along with D/iD and E/iE units in the CS/DS hybrid play important roles in the formation of the cerebellar neural network during postnatal brain development. << Less

  J. Biol. Chem. 281:18942-18952(2006) [PubMed] [EuropePMC]