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In this study we demonstrate that a disarmed version of the cytotoxin ricin can deliver exogenous CD8(+) T cell epitopes into the MHC class I-restricted pathway by a TAP-independent, signal peptidase-dependent pathway. Defined viral peptide epitopes genetically fused to the N terminus of an attenuated ricin A subunit (RTA) that was reassociated with its partner B subunit were able to reach the early secretory pathway of sensitive cells, including TAP-deficient cells. Successful processing and presentation by MHC class I proteins was not dependent on proteasome activity or on recycling of MHC class I proteins, but rather on a functional secretory pathway. Our results demonstrated a role for signal peptidase in the generation of peptide epitopes associated at the amino terminus of RTA. We showed, first, that potential signal peptide cleavage sites located toward the N terminus of RTA can be posttranslationally cleaved by signal peptidase and, second, that mutation of one of these sites led to a loss of peptide presentation. These results identify a novel MHC class I presentation pathway that exploits the ability of toxins to reach the lumen of the endoplasmic reticulum by retrograde transport, and suggest a role for endoplasmic reticulum signal peptidase in the processing and presentation of MHC class I peptides. Because TAP-negative cells can be sensitized for CTL killing following retrograde transport of toxin-linked peptides, application of these results has direct implications for the development of novel vaccination strategies.

Type

Journal article

Journal

J Immunol

Publication Date

01/07/2002

Volume

169

Pages

99 - 107

Keywords

Animals, Antigen Presentation, Antigens, Viral, Cytotoxicity Tests, Immunologic, Dogs, Endoplasmic Reticulum, Genetic Engineering, Glycoproteins, H-2 Antigens, Histocompatibility Antigen H-2D, Hydrolysis, Membrane Proteins, Mice, Mice, Inbred C57BL, Peptide Fragments, Protein Processing, Post-Translational, Protein Transport, Rabbits, Recombinant Fusion Proteins, Ricin, Serine Endopeptidases, Tumor Cells, Cultured, Viral Core Proteins, Viral Proteins