Molecular Cloning and Characterization of Chondroitin-4-O-sulfotransferase-3

Abstract

We have identified and characterized an N-acetylgalactosamine-4-O-sulfotransferase designated chondroitin-4-sulfotransferase-3 (C4ST-3) (GenBank TM accession number AY120869) based on its homology to HNK-1 sulfotransferase (HNK-1 ST). The cDNA predicts an open reading frame encoding a type II membrane protein of 341 amino acids with a 12-amino acid cytoplasmic domain and a 311-amino acid luminal domain containing a single potential N-linked glycosylation site. C4ST-3 has the greatest amino acid sequence identity when aligned with chondroitin-4-O-sulfotransferase 1 (C4ST-1) (45%) but also shows significant amino acid identity with chondroitin-4-O-sulfotransferase 2 (C4ST-2) (27%), dermatan-4-O-sulfotransferase 1 (29%), HNK-1 ST (26%), N-acetylgalactosamine-4-O-sulfotransferase 1 (26%), and N-acetylgalactosamine-4-Osulfotransferase 2 (23%). C4ST-3 transfers sulfate to the C-4 hydroxyl of ␤1,4-linked GalNAc that is substituted with a ␤-linked glucuronic acid at the C-3 hydroxyl. The open reading frame of C4ST-3 is encoded by three exons located on human chromosome 3q21.3. Northern blot analysis reveals a single 2.1-kilobase transcript. C4ST-3 message is expressed in adult liver and at lower levels in adult kidney, lymph nodes, and fetal liver. Although C4ST-3 and C4ST-1 have similar specificities, the highly restricted pattern of expression seen for C4ST-3 suggests that it has a different role than C4ST-1.We and others have recently reported the cloning and functional characterization of members of the HNK-1 family of sulfotransferases. These include human HNK-1 sulfotransferase (HNK-1 ST) 1 itself (1, 2), GalNAc-4-ST1 (3-5), GalNAc-4-ST2 (5, 6), dermatan-4-sulfotransferase-1 (D4ST-1) (7), chondroitin-4-sulfotransferase-1 (C4ST-1) (8, 9), and chondroitin-4-sulfotransferase-2 (C4ST-2) (8). With the exception of HNK-1 ST itself, which transfers sulfate to the C-3 hydroxyl of terminal glucuronic acid in the sequence GlcUA␤1,3Gal␤1,4GlcNAc-R to produce the HNK-1 epitope SO 4 -3-GlcUA␤1,3Gal␤1,4GlcNAc-R, the members of this family of sulfotransferases are all GalNAc-4-sulfotransferases. GalNAc-4-ST1 and GalNAc-4-ST2 both transfer sulfate to the C-4 hydroxyl of terminal ␤1,4-linked GalNAc on N-linked oligosaccharides such as those found on the glycoprotein hormones lutropin and thyrotropin (10, 11). Whereas C4ST-1, C4ST-2, and D4ST-1 are also GalNAc-4-O-sulfotransferases, they only transfer sulfate to the C-4 hydroxyl of internal ␤1,4-linked GalNAc moieties within the repeating disaccharide sequences found in chondroitin and dermatan (7-9). We now report the cloning of another member of this family of sulfotransferases. Like C4ST-1, this new sulfotransferase, chondroitin-4-sulfotransferase-3 (C4ST-3) transfers sulfate to the C-4 hydroxyl of ␤1,4-linked GalNAc that is flanked by GlcUA residues in chondroitin. In contrast to C4ST-1, C4ST-3 is labile at 37°C and has a restricted distribution, suggesting that it may have a unique biological role in vivo. EXPERIMENTAL PROCEDURES Molecular Cloning of a cDNA Encoding Human Chon...

Specificities of Three Distinct Human Chondroitin/Dermatan N-Acetylgalactosamine 4-O-Sulfotransferases Demonstrated Using Partially Desulfated Dermatan Sulfate as an Acceptor

“…It showed a significant homology to the HNK-1ST family members of human origin (C4ST-1 (31, 32), C4ST-2 (31), GalNAc4ST-1 (35,36,38), GalNAc4ST-2 (37, 38), and HNK-1ST (34)) and is the sixth family member. The other cDNA also shared significant sequence identities and structural similarities with the same family members and was identified as chondroitin 4-O-sulfotransferase-3 (C4ST-3) recently reported as the seventh member (55). As shown in Fig.…”

“…In this study, we investigated the acceptor specificities of these enzymes except for C4ST-3, which was phylogenetically closest to C4ST-1 but showed negligible catalytic activity toward the above mentioned substrates. Although the low activity was reportedly due to its thermal instability (55), specific substrates other than chondroitin may exist for C4ST-3 in view of its restricted expression profile in tissues such as liver and kidney (55).…”

Dermatan sulfotransferase Chst14/D4st1, but not chondroitin sulfotransferase Chst11/C4st1, regulates proliferation and neurogenesis of neural progenitor cells

“…To elucidate whether the ablation of Chst11 or Chst14 causes a dysregulation of the expression of other members of the HNK-1 sulfotransferase family in NSCs, differential gene expression of the HNK-1 sulfotransferase family members Chst8/GalNAc-4-st1 (Hiraoka et al, 2001;Okuda et al, 2000a;Okuda et al, 2003;Xia et al, 2000), Chst9/GalNAc-4-st2 (Hiraoka et al, 2001;Kang et al, 2001;Okuda et al, 2003), Chst10/HNK1-st (Bakker et al, 1997;Ong et al, 1998), Chst11/C4st1 (Hiraoka et al, 2000;Okuda et al, 2000b;Yamauchi et al, 2000), Chst12/C4st2 (Hiraoka et al, 2000), Chst13/C4st3 (Kang et al, 2002) and Chst14/D4st1 in Chst14 2/2 NSCs, the expression of Chst14 mRNA was reduced below the detection limit (Fig. 1B) A unique CS-DS structure containing CS-A, CS-D and DS patterns is recognized by the monoclonal antibody 473HD (Clement et al, 1998;Ito et al, 2005) as a marker for radial glial cells (Kabos et al, 2004).…”

Differential Roles of TwoN-Acetylgalactosaminyltransferases, CSGalNAcT-1, and a Novel Enzyme, CSGalNAcT-2

“…Sulfation of chondroitin is important to exhibit various biological functions. Sulfation at position C-4 of GalNAc is directed by chondroitin-4-O-sulfotransferase-1 (C4ST-1) (30), C4ST-2 (31), and C4ST-3 (32), that at C-6 of GalNAc by chondroitin-6-O-sulfotransferase-1 (C6ST-1) (33) and C6ST-2 (34), and that at C-2 of GlcA by uronyl 2-sulfotransferase (35) in the GalNAc-GlcA backbone. It is still unclear whether the sulfation occurs after the synthesis of the long chondroitin chain or the sulfation and GalNAc-GlcA polymerization occur simultaneously.…”