Initiation of galactosaminoglycan biosynthesis. Separate galactosylation and dephosphorylation pathways for phosphoxylosylated decorin protein and exogenous xyloside.
By using various radiolabelled precursors, glycosylation and phosphorylation of decorin in a rat fibroblast cell line was investigated in the presence of increasing concentrations of p-nitrophenyl-O-beta-d-xylopyranoside. Decorin core protein glycanation was suppressed to approximately 25% of the normal level in the presence of 2 mm and 3 mm xyloside. Glycans/saccharides were released from the core protein and size-separated by gel chromatography. The intracellular decorin obtained from cells treated with 2 mm xyloside was substituted with Xyl and also with Gal-Xyl and Gal-Gal-Xyl, but not with longer saccharides. Only the trisaccharide contained an almost fully phosphorylated Xyl. We conclude that galactosylation of endogenous, xylosylated decorin and exogenous xyloside probably follow separate pathways or that xylosides and early decorin glycoforms are kept separated. At the addition of the first glucuronic acid the two pathways seem to merge and dephosphorylation of decorin takes place. Xyloside-primed and secreted galactosaminoglycan chains produced simultanously retained phosphorylated Xyl. Inadequate dephosphorylation could be due to excess substrate or to a short transit.time. As shown previously [Moses, J., Oldberg, A., Eklund, E. & Fransson, L.-A. (1997) Eur. J. Biochem. 248, 767-774], brefeldin A-arrested decorin is substituted with the linkage-region extended with an undersulphated and incomplete galactosaminoglycan chain. In cells treated with this drug, xylosides were unable to prime galactosaminoglycan synthesis and unable to inhibit glycosylation and phosporylation of decorin.
Moses
J
J
Department of Cell and Molecular Biology, Faculty of Medicine, Lund University, Sweden. jonatan.moses @medkem.lu.se
Oldberg
A
A
Fransson
L A
LA
D000818:Animals; D020126:Brefeldin A / Q000494:pharmacology; D002460:Cell Line; D058575:Decorin; D016326:Extracellular Matrix Proteins; D005347:Fibroblasts / Q000378:metabolism; D006027:Glycosides / Q000378:metabolism*; D010766:Phosphorylation; D011134:Polysaccharides / Q000096:biosynthesis* / Q000378:metabolism; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology; D011509:Proteoglycans / Q000378:metabolism*; D051381:Rats
C546880:Dcn protein, rat; D058575:Decorin; D016326:Extracellular Matrix Proteins; D006027:Glycosides; D011134:Polysaccharides; D011500:Protein Synthesis Inhibitors; D011509:Proteoglycans; C010807:xylosides; D020126:Brefeldin A; C007815:galactosaminoglycan