Processing of HIV-1 envelope glycoprotein for class I-restricted recognition: Dependence on TAP1/2 and mechanisms for cytosolic localization

Processing of viral proteins for recognition by CTL involves degradation of the proteins in the cytosol of an infected cell followed by transport of t he resulting peptides into the endoplasmic reticulum (ER) by the TAP1/2 com plex, Uncertainty exists over the site of processing of viral envelope (env ) proteins since the extracellular domains of env proteins are not present in the cytosol where the class I Ag-processing pathway begins. Rather, the ectodomains of env proteins are cotranslationally translocated into the ER during biosynthesis. To analyze env protein processing, we used the herpes simplex virus protein ICP47 to block peptide transport by TAP1/2 and examin ed the effects of TAP blockade on the processing of the HIV-1 env protein. For the majority of env-specific CD8(+) CTL, the processing pathway require d TAP1/2-mediated transport of cytosolic peptides into the ER. To determine how env peptides are generated in the cytosol, we analyzed the processing of two TAP1/2-dependent epitopes containing N-linked glycosylation sites. I n each case, processing involved glycosylation-dependent posttranslational modification of asparagine residues to aspartic acid. These results are con sistent with cotranslational translocation of env into the ER, where glycos ylation occurs. This is followed by export of a fraction of the newly synth esized protein into the cytosol, where it is deglycosylated, with conversio n of the asparagines to aspartic acid residues, Following cytoplasmic prote olysis, env peptides are retransported by TAP1/2 into the ER, where associa tion with class I occurs. Thus, the env protein can enter the class I pathw ay through multiple distinct processing mechanisms.