Endogenously synthesized peptide with an endoplasmic reticulum signal sequence sensitizes antigen processing mutant cells to class I-restricted cell-mediated lysis.

Abstract

SummaryThe HLA-A2-positive human mutant cell line T2 is not lysed by influenza virus-specific HLA A2-restricted cytotoxic lymphocytes after virus infection . However, lysis does occur when cells are incubated with the antigenic influenza matrix protein-derived peptide M57-68 . To examine the nature of this defect, T2 cells were transfected with two different plasmids . One plasmid encoded the peptide M57-68, and the other encoded the same peptide preceded by an endoplasmic reticulum translocation signal sequence. Mutant T2 cells expressing the M57-68 peptide without the signal sequence were not susceptible to lysis by M57-68-specific HLAA2-restricted cytotoxic T lymphocytes, whereas T2 cells expressing the M57-68 peptide plus signal sequence were lysed effectively. Lysis of parental T1 cells with either plasmid was equally effective . These results suggest that the T2 mutant cells are defective in the transport of antigenic peptides from the cytosol into the secretory pathway.

Cutting Edge: Efficient MHC Class I Cross-Presentation during Early Vaccinia Infection Requires the Transfer of Proteasomal Intermediates between Antigen Donor and Presenting Cells

“…To determine whether cross-presentation of the mature epitope could be detected by increasing its chances to bind gp96 or other ER resident chaperones, we expressed 258 -265 directly in the ER of donor cells using a plasmid encoding (SS)258 -265 where (SS) is the ER insertion/signal sequence derived from the adenovirus E3/19K glycoprotein that is removed cotranslationally (19). However, we did not detect cross-presentation of (SS)258 -265 (Fig.…”

Generation of MHC Class I Peptide Antigens by Protein Processing in the Secretory Route by Furin

“…Also, peptide translocation by TAP is not a necessary requisite for class I loading, as peptides present in the ER may bind to MHC class I, either directly or by the cooperation of chaperones. Therefore, any method of introducing class I-fitting complementary peptides in the ER in sufficient amounts may result in binding and presentation by class I molecules, as demonstrated by the high efficiency of presentation of ER-targeted minigene products (6). Similarly, any proteolytic activity, besides the proteasome, that is able to generate short peptides either in the cytosol or inside the ER would be suitable for antigen presentation.…”

Potent Cytolytic Response by a CD8+ CTL Clone to Multiple Peptides from the Same Protein in Association with an Allogeneic Class I MHC Molecule

“…That the presumed expression of p2Ca in the ER of transfected C1R-L d cells did not render the cells susceptible to lysis by 2C CTL (see Table I) is at odds with experiments demonstrating that optimal peptides expressed in the ER are usually effectively presented on the cell surface (14,15). A low copy number of this minigene in the transfected cells may also account for the lack of detectable peptide presentation.…”

“…C1R-L d and T2-L d cells were transfected with p8901 plasmids encoding p2Ca or p2Cb or their variants as well as with a plasmid containing the adenovirus E3/19-kDa protein ER translocation signal sequence RYMIL GLLALAAVCSA(A) followed by the QL9 sequence as previously described (13)(14)(15). The transfected cells are referred to as C1R-L d (p2Ca), C1R-L d (p2Cb), and C1R-L d (sig-QL9), respectively (see Table I).…”