Chondroitin 4-sulfotransferase (C4ST) catalyzes the transfer of sulfate from 3-phosphoadenosine 5-phosphosulfate to position 4 of N-acetylgalactosamine residue of chondroitin. The enzyme has been previously purified to apparent homogeneity from the serum-free culture medium of rat chondrosarcoma cells (Yamauchi, A., Hirahara, Y., Usui, H., Takeda, Y., Hoshino, M., Fukuta, M., Kimura, J. H., and Habuchi, O. (1999) J. Biol. Chem. 274, 2456 -2463). The purified enzyme also catalyzed the sulfation of partially desulfated dermatan sulfate. We have now cloned the cDNA of the mouse C4ST on the basis of the amino acid sequences of peptides obtained from the purified enzyme by protease digestion. This cDNA contains a single open reading frame that predicts a protein composed of 352 amino acid residues. The protein predicts a Type II transmembrane topology. The predicted sequence of the protein contains all of the known amino acid sequence and four potential sites for N-glycosylation, which corresponds to the observation that the purified C4ST is an N-linked glycoprotein. The amino acid sequence of mouse C4ST showed significant sequence homology to HNK-1 sulfotransferase. Comparison of the sequence of mouse C4ST with human HNK-1 sulfotransferase revealed ϳ29% identity and ϳ48% similarity at the amino acid level. When the cDNA was introduced in a eukaryotic expression vector and transfected in COS-7 cells, the sulfotransferase activity that catalyzes the transfer of sulfate to position 4 of GalNAc residue of both chondroitin and desulfated dermatan sulfate was overexpressed. Northern blot analysis showed that, among various mouse adult tissues, 5.7-kilobase message of C4ST was mainly expressed in the brain and kidney.Chondroitin sulfate proteoglycans are found in various tissues as molecules having divergent molecular architecture (1, 2). Chondroitin sulfate chains attached to chondroitin sulfate proteoglycans appear to play important roles in the formation and maintenance of cartilage tissue, because undersulfation of chondroitin sulfate resulted from the defective synthesis of PAPS 1 (3, 4) or defective sulfate transport (5) was found to cause underdevelopment of skeleton. Various chondroitin sulfate proteoglycans have been reported to be present in the brain (6, 7) and to function in the regulation of neurite outgrowth and neural cell adhesion (8 -12), neuronal migration (13), and the survival of neurons (14). Chondroitin sulfate chains are also shown to be involved in the interaction with CD44 (15, 16), phospholipase A 2 (17), Plasmodium falciparuminfected erythrocytes (18), and L-selectin (19). Chondroitin sulfates have sulfate group at various positions of the component sugars; position 6 and/or 4 of GalNAc residues and position 2 or 3 of GlcA residues. The pattern of sulfation of chondroitin sulfate chains varies with the source of the proteoglycans, development of animal (20 -22), and malignant change (23), suggesting that sulfate moieties attached to the specific position of the component sugars may be related to ...

Chondroitin 4-sulfotransferase (C4ST) catalyzes the transfer of sulfate from 3'-phosphoadenosine 5'-phosphosulfate to position 4 of the N-acetylgalactosamine residues of chondroitin. We previously reported the cloning of C4ST cDNA from mouse brain. We here report the cloning and expression of human C4ST cDNA. The cDNA was isolated from a human fetal brain cDNA library by hybridization with a DNA probe prepared from rat poly(A)(+) RNA used for the cloning of mouse C4ST cDNA. The cDNA comprises a single open reading frame that predicts a Type II transmembrane protein composed of 352 amino acids. The protein has an amino acid sequence homology of 96% with mouse C4ST. When the cDNA was introduced into a eukaryotic expression vector and transfected in COS-7 cells, the sulfotransferase activity that transfers sulfate to both chondroitin and desulfated dermatan sulfate was overexpressed. Northern blot analysis indicated that human C4ST mRNAs (6.0 and 1.9 kb) are expressed ubiquitously in various adult human tissues. Dot blot analysis has shown that human C4ST is strongly expressed in colorectal adenocarcinoma and peripheral blood leukocytes, whereas strong expression of human chondroitin 6-sulfotransferase (C6ST) is observed in aorta and testis. These observations suggest that the expression of C4ST and C6ST may be controlled differently in human tissues. The C4ST gene was localized to chromosome 12q23.2-q23.3 by fluorescence in situ hybridization.