Secretion of high-mannose-type alpha 1-proteinase inhibitor and alpha 1-acid glycoprotein by primary cultures of rat hepatocytes in the presence of the mannosidase I inhibitor 1-deoxymannojirimycin.
Two different forms of alpha 1-proteinase inhibitor and alpha 1-acid glycoprotein were found in primary cultures of rat hepatocytes. After a 2.5-h labeling period with [35S]methionine the high-mannose-type precursor of alpha 1-proteinase inhibitor (Mr 49000) and alpha 1-acid glycoprotein (Mr 39 000) and the mature-complex-type alpha 1-proteinase inhibitor (Mr 54 000) and alpha 1-acid glycoprotein (Mr 43 000-60 000) could be immunoprecipitated from the cells, but only the complex-type forms of the two glycoproteins were secreted into the hepatocyte media. When hepatocytes were incubated with the mannosidase I inhibitor 1-deoxymannojirimycin at a concentration of 4 mM, the 49 000-Mr form of alpha 1-proteinase inhibitor and the 39 000-Mr form of alpha 1-acid glycoprotein could be detected in the cells as well as in their media. Neither the secretion of alpha 1-proteinase inhibitor nor that of alpha 1-acid glycoprotein was impaired by 1-deoxymannojirimycin. While alpha 1-proteinase inhibitor and alpha 1-acid glycoprotein, secreted by control cells, were resistant to endoglucosaminidase H, alpha 1-proteinase inhibitor and alpha 1-acid glycoprotein, secreted by hepatocytes treated with 4 mM 1-deoxymannojirimycin, could be deglycosylated by endoglucosaminidase H. When the [3H]mannose-labeled oligosaccharides of alpha 1-proteinase inhibitor, secreted by 1-deoxymannojirimycin-treated hepatocytes, were cleaved off by endoglucosaminidase H and analyzed by Bio-Gel P-4 chromatography, they eluted at the position of Man9GlcNAc, indicating that mannosidase I had been efficiently inhibited. 1-Deoxymannojirimycin did not inhibit the synthesis or the cotranslational N-glycosylation of alpha 1-proteinase inhibitor or alpha 1-acid glycoprotein.
Gross
V
V
Steube
K
K
Tran-Thi
T A
TA
McDowell
W
W
Schwarz
R T
RT
Decker
K
K
Gerok
W
W
Heinrich
P C
PC
D017485:1-Deoxynojirimycin; D000180:Acrylic Resins; D000818:Animals; D001798:Blood Proteins / Q000096:biosynthesis / Q000378:metabolism*; D002478:Cells, Cultured; D055598:Chemical Phenomena; D011232:Chemical Precipitation; D002621:Chemistry; D002845:Chromatography / Q000379:methods; D005944:Glucosamine / Q000031:analogs & derivatives / Q000494:pharmacology; D006596:Hexosaminidases; D007120:Immunochemistry; D008099:Liver / Q000187:drug effects / Q000378:metabolism*; D008358:Mannose; D008361:Mannosidases / Q000037:antagonists & inhibitors*; D017038:Mannosyl-Glycoprotein Endo-beta-N-Acetylglucosaminidase; D009844:Oligosaccharides / Q000032:analysis; D009961:Orosomucoid / Q000096:biosynthesis / Q000378:metabolism*; D051381:Rats; D000515:alpha 1-Antitrypsin
D000180:Acrylic Resins; D001798:Blood Proteins; D009844:Oligosaccharides; D009961:Orosomucoid; D000515:alpha 1-Antitrypsin; D017485:1-Deoxynojirimycin; C046089:Bio-Gel P-4; D006596:Hexosaminidases; D008361:Mannosidases; C046090:mannosyl-oligosaccharide 1,2-alpha-mannosidase; D017038:Mannosyl-Glycoprotein Endo-beta-N-Acetylglucosaminidase; D005944:Glucosamine; D008358:Mannose