Molecular cloning and expression of two distinct human chondroitin 4-O-sulfotransferases that belong to the HNK-1 sulfotransferase gene family.
Using an expression cloning strategy, the cDNA encoding the human HNK-1 sulfotransferase (HNK-1ST) has been cloned. During this cloning we found that HNK-1ST and other Golgi-associated sulfotransferases cloned before share homologous sequences including the RDP motif (Ong, E., Yeh, J.-C., Ding, Y., Hindsgaul, O., and Fukuda, M. (1998) J. Biol. Chem. 223, 5190-5195). Using this conserved sequence in HNK-1ST as a probe, we identified two expressed sequence tags in EST data base which have 31.6 and 30.7% identity with HNK-1ST at the amino acid levels. Expression of these two full-length cDNAs failed to form HNK-1 glycan nor to add sulfate to CD34 or NCAM. Surprisingly, proteins expressed by these cDNAs transferred sulfate to the C-4 position of N-acetylgalactosamine in chondroitin and desulfated dermatan sulfate, thus we named these two enzymes, chondroitin 4-O-sulfotransferase 1 and -2 (C4ST-1 and C4ST-2). Both C4ST-1 and C4ST-2, however, did not form 4, 6-di-O-sulfated N-acetylgalactosamine when chondroitin sulfate C was used as an acceptor. Moreover, analysis of (35)S-labeled dermatan sulfate formed by C4ST-1 indicate that sulfation preferentially took place in GlcA-->GalNAc unit than in IdoA-->GalNAc unit, suggesting that 4-O-sulfation at N-acetylgalactosamine may precede epimerization of glucuronic acid to iduronic acid during dermatan sulfate biosynthesis. Northern analysis demonstrated that the transcript for C4ST-1 is predominantly expressed in peripheral leukocytes and hematopoietic tissues while the C4ST-2 transcript is more widely expressed in various tissues. These results indicate C4ST-1 and C4ST-2 play complementary roles in chondroitin and dermatan sulfate synthesis in different tissues.
Hiraoka
N
N
Glycobiology Program, Cancer Research Center, The Burnham Institute, La Jolla, California 92037, USA.
Nakagawa
H
H
Ong
E
E
Akama
T O
TO
Fukuda
M N
MN
Fukuda
M
M
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D015152:Blotting, Northern; D016466:CHO Cells; D002851:Chromatography, High Pressure Liquid; D002874:Chromosome Mapping; D003001:Cloning, Molecular; D006224:Cricetinae; D018076:DNA, Complementary / Q000378:metabolism; D016208:Databases, Factual; D003871:Dermatan Sulfate / Q000378:metabolism; D020224:Expressed Sequence Tags; D006031:Glycosylation; D006801:Humans; D015151:Immunoblotting; D008969:Molecular Sequence Data; D010802:Phylogeny; D020033:Protein Isoforms; D017386:Sequence Homology, Amino Acid; D015238:Sulfotransferases / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D013997:Time Factors; D014018:Tissue Distribution; D014162:Transfection; D000097811:Carbohydrate Sulfotransferases
D018076:DNA, Complementary; D003871:Dermatan Sulfate; D020033:Protein Isoforms; D015238:Sulfotransferases; C499544:CHST13 protein, human; C411659:CHST11 protein, human; C411660:CHST12 protein, human; C499549:CHST10 protein, human; D000097811:Carbohydrate Sulfotransferases; C109318:HNK-1 sulfotransferase; C019893:chondroitin 4-sulfotransferase