Targeting and processing of glycophorins in murine erythroleukemia cells: use of brefeldin A as a perturbant of intracellular traffic.
We previously showed that glycophorins are expressed in virus-transformed, murine erythroleukemia cells; we detected four glycophorin precursors (two more than in normal erythroblasts) and found that two of them are not translocated or are inefficiently translocated across the endoplasmic reticulum (ER) membrane. By using the drug brefeldin A to block intracellular transport of proteins from the ER to the Golgi complex, the translocated precursors were shown to accumulate in the ER, while the untranslocated forms were rapidly degraded with an intracellular half-life of approximately 20 min. Brefeldin A did not inhibit the synthesis of fatty acylation of the precursors but substantially delayed their acquisition of O-linked oligosaccharides, which indicates that murine glycophorins are fatty acylated in the ER and O-glycosylated in the Golgi complex. Even after 6 hr in brefeldin A, glycophorins were only partially glycosylated, resulting in the accumulation of glycoproteins apparently sialylated but lower in apparent molecular mass than mature glycophorins. Complete glycophorin processing resumed only after removal of the drug. In murine erythroleukemia cells, brefeldin A caused a rapid and extensive disorganization of the entire Golgi complex accompanied by the accumulation of membranes in a part of the ER closely associated with ER transitional elements. These findings extend recently published results [Lippincott-Schwartz, J., Yuan, L. C., Bonifacino, J. S. & Klausner, R. D. (1989) Cell 56, 801-813] and suggest that brefeldin A induces net membrane flow from the entire Golgi complex to the ER.
Department of Cell Biology, Yale University School of Medicine, New Haven, CT 06510.
J B
JB
Ulmer
G E
GE
Palade
D000818:Animals; D000935:Antifungal Agents / Q000494:pharmacology; D020126:Brefeldin A; D002460:Cell Line; D003517:Cyclopentanes / Q000494:pharmacology*; D006021:Glycophorins / Q000096:biosynthesis* / Q000235:genetics / Q000302:isolation & purification; D006031:Glycosylation; D007700:Kinetics; D004915:Leukemia, Erythroblastic, Acute / Q000378:metabolism*; D051379:Mice; D008854:Microscopy, Electron; D009363:Neoplasm Proteins / Q000096:biosynthesis / Q000302:isolation & purification; D011499:Protein Processing, Post-Translational; D012795:Sialoglycoproteins / Q000096:biosynthesis*
D000935:Antifungal Agents; D003517:Cyclopentanes; D006021:Glycophorins; D009363:Neoplasm Proteins; D012795:Sialoglycoproteins; D020126:Brefeldin A