Abnormal glycosylation of human fibronectin secreted in the presence of monensin.
Detailed studies of the effects of the ionophore monensin upon the glycosylation of secreted fibronectin have been carried out. Human fibroblasts in culture were incubated in 1 microM monensin for several hours, following which radiolabeled glucosamine or mannose was added to the cultures. Parallel incubation and labeling of control cultures were done. Labeled fibronectin was isolated from the culture media by gelatin-Sepharose chromatography, from cell surfaces by urea extraction, and from intracellular locations by cell lysis followed by immunoprecipitation. Detailed comparison of the glycopeptides released from fibronectin by pronase and of the oligosaccharides liberated by hydrazinolysis was carried out, particularly focusing on the secreted fibronectin, using gel filtration, high performance liquid chromatography, and concanavalin A chromatography, in conjunction with the use of endoglycosidase H and specific exoglycosidases. We demonstrate that fibronectin in the medium of monensin-treated cultures differs in its glycosylation pattern from the control fibronectin. High mannose oligosaccharides are abundant in the monensin-derived fibronectin, whereas the control protein contains primarily complex oligosaccharides. Monensin apparently does not alter the initial glycosylation of fibronectin since the high mannose oligosaccharides are present on both control and monensin-treated intracellular fibronectin. We suggest, therefore, that monensin, by impairing intracellular translocation through the Golgi region, allows incompletely processed forms of fibronectin to reach the cell surface and to be released into the culture medium.
D000328:Adult; D002478:Cells, Cultured; D005347:Fibroblasts / Q000187:drug effects / Q000378:metabolism; D005353:Fibronectins / Q000302:isolation & purification / Q000378:metabolism*; D005663:Furans / Q000494:pharmacology*; D005944:Glucosamine / Q000378:metabolism*; D006020:Glycopeptides / Q000032:analysis; D006801:Humans; D007700:Kinetics; D008358:Mannose / Q000378:metabolism*; D008985:Monensin / Q000494:pharmacology*; D009844:Oligosaccharides / Q000032:analysis; D012867:Skin / Q000378:metabolism*
D005353:Fibronectins; D005663:Furans; D006020:Glycopeptides; D009844:Oligosaccharides; D008985:Monensin; D005944:Glucosamine; D008358:Mannose