TRANSPORT OF A LYSOSOMALLY TARGETED ROUS-SARCOMA VIRUS ENVELOPE GLYCOPROTEIN INVOLVES TRANSIENT EXPRESSION ON THE CELL-SURFACE

The details of intracellular transport pathways for glycosylated prote ins remain incompletely described. We previously described a mutant Ro us sarcoma virus envelope glycoprotein (gp), mu 26, with an altered me mbrane-spanning domain that was targeted to lysosomes after traversing the trans-Golgi. This mutant protein was not detectable on the cell s urface by immunofluorescence, but its pathway for degradation remained unclear. To investigate this we have employed a second env mutation, sig, that results in a protein which is defective for normal cleavage/ activation by intracellular enzymes, but remains susceptible to cleava ge by extracellular proteases. Cleavage/activation of the double mutan t by trypsin, which could only occur if it was exposed on the cell sur face, was observed, indicating that the plasma membrane is an intermed iate destination in the transport of this mutant protein. To substanti ate these results, cells expressing the mu 26 glycoprotein were incuba ted with an antibody specific for the native protein in the presence o f chloroquine. The specific accumulation of this antibody/gp complex i n vesicles, as detected by internal immunofluorescence, confirmed the trypsin cleavage results. We conclude that this rapidly degraded mutan t protein is transported from the trans-Golgi to the cell surface, whe re it is only transiently exposed, and then rapidly endocytosed and ly sosomally degraded. The relevance of these results to the targeting of lysosomal proteins is discussed. (C) 1995 Academic Press, Inc.