Intracellular degradation of the HIV-1 envelope glycoprotein - Evidence for, and some characteristics of, an endoplasmic reticulum degradation pathway

Analysis of the fate of HIV-1 envelope protein gp160 (Env) has shown that n ewly synthesized proteins may he degraded within the biosynthetic pathway a nd that this degradation may take place in compartments Ether than the lyso somes. The fate of newly synthesized Em was studied in living BHK-21 cells with the recombinant vaccinia virus expression system. We found that gp160 not only undergoes physiological endoproteolytic cleavage, producing gp120, but is also degraded. producing proteolytic fragments of 120 kDa to 26 kDs in size, as determined by SDS/PAGE in non reducing conditions. Analysis of the 120-kDa proteolytic fragment, and comparison with gp120, showed that i t is composed of peptides linked by disulfides bonds and lacks the V3-loop epitope and the C-terminal domain of gp120 (amino acids 506-516). A permeab ilized cell system, with impaired transport of labeled Env from the endopla smic reticulum (ER) to Golgi compartments, was developed to determine the s ite of degradation and to define some biochemical characteristics of the in tracellular degradation process. In the semipermeable BHK-21 cells, there w as: (a) no gp 120 production (b), a progressive decrease in the amount of n ewly synthesized gp160 and a concomittant increase in the amount of a 120-k Da protelolytic fragment. This fragment had the same biochemical characteri stics as the 120-kDa proteolytic fragment found in living nonpermeabilized cells, and (c) susceptibility of the V3 loop. This degradation process occu rred in the ER, as shown by both biochemical and indirect immunofluorescenc e analysis. Futhermore, there was evidence that changes in redox state are involved in the ER-dependmt envelope degradation pathway because adding red ucing agents to permeabilized cells caused dose-dependent degradation of th e 120-kDa proteolytic fragment and of the remaining gp160 glycoprotein. Thu s our results provide direct evidence that regulated degradation of the HIV -1 envelope glycoprotein may take piece in the ER of infected cells.