Reduced temperature prevents transfer of a membrane glycoprotein to the cell surface but does not prevent terminal glycosylation

Abstract

The transport kinetics of the influenza virus hemagglutinin from its site of synthesis to the apical plasma membrane of Madin-Darby canine kidney cells, a polarized epithelial cell line, were studied by a sensitive tryptic assay. Hemagglutinin acquired terminal sugars, as judged by sensitivity to endo-beta-N-acetylglucosaminidase H, 10-15 min after synthesis, and first appeared on the apical domain 15 min later. None of the pulse-labeled hemagglutinin accumulated on the basolateral domain. At 20 degrees C, terminal glycosylation continued, but no hemagglutinin was detected on the cell surface within 2 hr. If the incubation temperature was raised from 20 degrees C to 37 degrees C, hemagglutinin was quickly externalized, demonstrating that the inhibition at low temperature was reversible.