Effect of glycosidase inhibitors on the biosynthesis of alpha 2-plasmin inhibitor and antithrombin III in Hep G2 cells.
We studied the effect of the glucosidase I inhibitor, N-methyl-1-deoxynojirimycin (MdN) and the mannosidase inhibitor, 1-deoxymannojirimycin (dMM) on the biosynthesis and secretion of alpha 2-plasmin inhibitor (alpha 2-PI) and antithrombin III (ATIII) in cultures of human hepatoma (Hep-G2) cells. Incubation with 1 mM MdN decreased secreted alpha 2-PI activity and antigen levels by about 40%, whereas those of ATIII were not affected. Neither inhibitor affected the messenger RNA levels as determined by Northern blotting. Pulse-chase studies using [35S]-methionine showed that MdN decreased alpha 2-PI and ATIII secretion rates. By the 18 h chase, MdN had decreased secreted alpha 2-PI to 50-60%, with little effect on ATIII. Intracellular forms of alpha 2-PI or ATIII synthesized by cells treated with 1 mM MdN were sensitive to endoglycosidase H (Endo H), whereas almost all the secreted forms were resistant, suggesting the presence of complex-type oligosaccharides. In the presence of 1 mM dMM, cells synthesized Endo H-sensitive alpha 2-PI and ATIII with similar secretion rates. These results suggest that retention of glucose on N-linked oligosaccharides not only retards the exit of alpha 2-PI and ATIII, but also changes the catabolic rate of alpha 2-PI in the endoplasmic reticulum.
Mori
K
K
Division of Thrombosis and Hemostasis, Jichi Medical School, Tochigi-ken, Japan.
Wada
Y
Y
Mimuro
J
J
Matsuda
M
M
Yoshikuni
Y
Y
Kimura
K
K
Sakata
Y
Y
D017485:1-Deoxynojirimycin / Q000031:analogs & derivatives* / Q000494:pharmacology*; D000818:Animals; D000990:Antithrombin III / Q000096:biosynthesis*; D004285:Dogs; D006026:Glycoside Hydrolases / Q000037:antagonists & inhibitors* / Q000235:genetics; D006596:Hexosaminidases; D006801:Humans; D008361:Mannosidases / Q000037:antagonists & inhibitors / Q000235:genetics; D017038:Mannosyl-Glycoprotein Endo-beta-N-Acetylglucosaminidase; D012333:RNA, Messenger / Q000032:analysis; D014407:Tumor Cells, Cultured / Q000378:metabolism; D000979:alpha-2-Antiplasmin / Q000096:biosynthesis*
D012333:RNA, Messenger; D000979:alpha-2-Antiplasmin; D017485:1-Deoxynojirimycin; D000990:Antithrombin III; D006026:Glycoside Hydrolases; D006596:Hexosaminidases; D008361:Mannosidases; D017038:Mannosyl-Glycoprotein Endo-beta-N-Acetylglucosaminidase; C033424:N-methyldeoxynojirimycin