PROCESSING OF VIRAL ENVELOPE GLYCOPROTEIN BY THE ENDOMANNOSIDASE PATHWAY - EVALUATION OF HOST-CELL SPECIFICITY

Endo-alpha-D-mannosidase is an enzyme involved in N-linked oligosaccha ride processing which through its capacity to cleave the internal link age between the glucose-substituted mannose and the remainder of the p olymannose carbohydrate unit can provide an alternate pathway for achi eving deglucosylation and thereby make possible the continued formatio n of complex oligosaccharides during a glucosidase blockade. In view o f the important role which has been attributed to glucose on nascent g lycoproteins as a regulator of a number of biological events, we chose to further define the in vivo action of endomannosidase by focusing o n the well characterized VSV envelope glycoprotein (G protein) which c an be formed by the large array of cell lines susceptible to infection by this pathogen, Through an assessment of the extent to which the G protein was converted to an endo-beta-N-acetylglucosaminidase (endo H) -resistant form during a castanospermine imposed glucosidase blockade, we found that utilization of the endomannosidase-mediated deglucosyla tion route was clearly host cell specific, ranging from greater than 9 0% in HepG2 and PtK1 cells to complete absence in CHO, MDCK, and MDBK cells, with intermediate values in BHK, BW5147,3, LLC-PK1, BRL, and NR K cell lines. In some of the latter group the electrophoretic pattern after endo H treatment suggested that only one of the two N-linked oli gosaccharides of the G protein was processed by endomannosidase, In th e presence of the specific endomannosidase inhibitor, Glc alpha 1 --> 3(1-deoxy)mannojirimycin, the conversion of the G protein into an endo H-resistant form was completely arrested. While the lack of G protein processing by CHO cells was consistent with the absence of in vitro m easured endomannosidase activity in this cell line, the failure of MDB K and MDCK cells to convert the G protein into an endo H-resistant for m was surprising since these cell lines have substantial levels of the enzyme. Similarly, we observed that influenza virus hemagglutinin was not processed in castanospermine-treated MDCK cells. Our findings sug gest that studies which rely on glucosidase inhibition to explore the function of glucose in controlling such critical biological phenomena as intracellular movement or quality control should be carried out in cell lines in which the glycoprotein under study is not a substrate fo r endomannosidase action.