The function of the interferon-γ-inducible subunits and activators of the proteasome during the immuneresponse against the lymphocytic chriomeningitis virus

The presentation of viral antigens on MHC class I molecules requires their intracellular fragmentation into peptides of appropriate length and anchor residue positions. Evidence has accumulated that the proteasome is the endoprotease in charge of the generation of MHC class I ligands in the cytoplasm. The generation of T cell epitopes derived from the leader peptides of endoplasmic reticulum (ER) targeted proteins, however, has been reported to be independent of the proteasome. Here we show that the H-2DD restricted antigen presentation of the immunodominant T cell epitope derived from the ER leader of the glycoprotein of lymphocytic choriomeningitis virus (LCMV) is completely abolished by administration of the proteasome inhibitor lactacystin. Thus our data support the role of the proteasome in class 1 restricted antigen processing and extend it to an ER leader derived epitope from a viral glycoprotein. Introduction Two ways have been proposed how peptide ligands for MHC class I molecules can be generated. The first and quantitatively more prominent way is the endoproteolytic fragmentation in the cytoplasm followed by delivery of peptides via the transporter associated with antigen presentation (TAP) into the ER. The proteasome has been implicated in the generation of these peptide ligands for a number of reasons (reviewed in Groettrup et al, 1996). The demonstration that peptide aldehyde inhibitors of the proteasome like LLnL strongly inhibit peptide dependent MHC class I assembly and maturation as well as antigen presentation (Rock et al, 1994; Hughes et al., 1996) has recently been corroborated by further experiments employing the most specific proteasome inhibitor available to date: lactacystin (Craiu et al, 1997; Cerundolo et al, 1997). Lactacystin is a metabolite from Streptomyces (Omura et al, 1991) which upon formation of the clasto-lactacystin ß-lactone inhibits proteasomes by covalent modification of its active site threonine residues (Dick et al, 1996; Fenteany et al, 1995). The second way of peptide generation for class I molecules has been reported to be independent of proteasome activity (Henderson et al, 1992; Wei and Cresswell, 1992; Hughes, et al., 1996; Cerundolo, et al., 1997). The source of these MFIC ligands are mainly leader sequences of ER targeted proteins and they are loaded onto class I molecules independent of TAP transporters. Their location within an ER signal sequence and the processing of leader sequences during cotranslational insertion appears to be sufficient for ER targeting and allows further processing in the ER. However, Lyko et ai. have shown that cleaved signal peptides may be further processed and that resulting fragments can be released into the cytosol (Lyko et al, 1995; Martoglio et al, 1997). This situation may apply to the four examples known to date where leader derived MHC ligands are presented in a TAP dependent manner: 1) The first example was reported by Hombach et al. who have shown that the immunodominant II-2DD restricted epitope gp33 from the LCMV glycoprotein ER leader is presented in a strictly TAP dependent manner (Hombach et al, 1995). 2) For the Friend, Moloney, and Rauscher strain of Murine Leukemia Virus an immunodominant and 11-20^ restricted T cell epitope has recently been identified