O-glycosylation of intact and truncated ribophorins in brefeldin A- treated cells: newly synthesized intact ribophorins are only transiently accessible to the relocated glycosyltransferases

Subconlluent cultures of HeLa-RI332 cells were incubated with fresh media with or without 5/~g/rnl BFA at 370C for 2 h and fixed in 2% glutaraldehyde in 0.1 M sodium cacodylate buffer, pH 7.4, for 45 vain on ice. The cells were then scrap~ off the dish, sedimented, postfixed with glutaraldehyde and OsO4, and processed for routine EM. Thin sections were stained with uranyl acetate and lead citrate and examined in an electron micro= SCOpe operated at 80 kV (model 301; Philips Electronic Instruments Co., Mahwah, NJ).