Peptide transporter‐independent, stress protein‐mediated endosomal processing of endogenous protein antigens for major histocompatibility complex class I presentation

Abstract

The peptide transporter-defective cell line RMA-S expressing the wild-type simian virus 40 large T antigen (wtT-Ag) from a transfected gene did not present two well-defined, H-2 class I (Db)-restricted epitopes of T-Ag to cytotoxic T lymphocytes (CTL). Hence, "endogenous" processing and presentation of the wtT-Ag depended on a functional peptide transporter heterodimer. In contrast, both T-Ag epitopes were efficiently presented to CTL by transfected RMA-S cells expressing a truncated, cytoplasmic T-Ag variant (cT-Ag) or a karyophilic, amino-terminal 272-amino acid T-Ag fragment. Transporter-independent "endogenous" processing of mutant T-Ag molecules correlated with their association with the constitutively expressed heat shock protein 73 (hsp73). Class I-restricted presentation of both epitopes processed from these hsp73-associated protein antigens was sensitive to NH4Cl and chloroquine. These data indicate that selected intracellular proteins access an alternative, hsp73-mediated pathway for class I-restricted presentation that operates independent of peptide transporters in an endosomal compartment.

Enhanced Priming of Multispecific, Murine CD8+ T Cell Responses by DNA Vaccines Expressing Stress Protein-Binding Polytope Peptides

“…4). This confirms that the tight, noncovalent association between hsp73 and the viral DnaJ domain requires the intact 77-residue domain as described by us in other Ag systems (10,11,13,15,16). Hence, we have constructed DNA vaccines carrying the immunogenic pt10 domain, but differing in the level of expression and the association with hsp73.…”

“…In addition to facilitating expression, hsp binding offers other features that make it attractive as a vaccine. Hsp molecules of the hsp70 and hsp90 class are intrinsic adjuvants (as indicated above) and introduce cytosolic proteins into alternative processing pathways (10,11). As different proteolytic systems seem to give rise to different, although overlapping, repertoires of antigenic epitopes (49), this means of expression can potentially extend the repertoire of immunogenic epitopes presented by an Ag.…”

Restoration of MHC Class I Surface Expression and Endogenous Antigen Presentation by a Molecular Chaperone

“…in binding unfolded proteins and peptides and in mediating their translocation across lipid bilayers [30], as well as in enhancing the formation of multimeric protein complexes [8]. This role is supported further by the recent results of Schirmbeck & Reimann [31], Suto & Srivastava [32] and Arnold et al [33], which indicate that the entry of antigenic peptides into the MHC class I pathway can be mediated directly by a molecular chaperone.…”

Priming Protective CD8 T Cell Immunity by DNA Vaccines Encoding Chimeric, Stress Protein-Capturing Tumor-Associated Antigen

“…Levels of immunoprecipitated proteins were analyzed by fluorography of the gels. Alternatively, nonlabeled cell extracts were immunoprecipitated as described above and processed for hsp73-or T-Ag-specific Western blotting as described previously (22,23). Briefly, immunoprecipitates were processed by SDS-PAGE and blotted onto nitrocellulose paper.…”