Intracellular processing and presentation of T cell epitopes, expressed by recombinant Escherichia coli and Salmonella typhimurium, to human T cells

Abstract

Vaccines based on recombinant attenuated bacteria represent a potentially safe and effective immunization strategy. A carrier system was developed to analyze in vitro whether foreign T cell epitopes, inserted in the outer membrane protein PhoE of Escherichia coli and expressed by recombinant bacteria, are efficiently processed and presented via human leukocyte antigen (HLA) class I and II molecules by bacterial infected human macrophages. A well-defined HLA-B27-restricted cytotoxic T cell (CTL) epitope and an HLA-DR53 restricted T helper (Th) epitope of the fusion protein of measles virus were genetically inserted in a surface-exposed region of PhoE, and the chimeric proteins were expressed in E. coli and Salmonella typhimurium. Macrophages infected with both recombinant bacteria presented the Th epitope to the specific CD4+ T cell clone, but failed to present the CTL epitope to the specific CD8+ T cell clone. Presentation of the Th epitope by the infected macrophages was inhibited by cytochalasin D, indicating that phagocytic processing of intact bacteria within infected macrophages was essential for antigen presentation via HLA class II. Presentation of the Th epitope to the CD4+ T cell clone by infected macrophages was blocked by brefeldin A and cycloheximide, indicating the requirement of nascent HLA class II molecules for presentation. The efficiency of macrophages to process and present the inserted Th epitope was similar for both the recombinant E. coli and S. typhimurium strains.

Identification of a Common HLA-DP4-Restricted T-Cell Epitope in the Conserved Region of the Respiratory Syncytial Virus G Protein

“…TCC 1 was generated from nasalbrush cells collected from an infant (age, 20 months) during the acute phase of a laboratory-confirmed RSV-mediated upper respiratory tract infection. The nasal-brush T cells were stimulated in vitro with autologous ␥-irradiated (3,000 rad) BLCL-RSV, and TCCs were generated by limiting dilution as described before (35). In short, T cells were seeded in 60-well Terasaki plates (Greiner Bio-One, Frickenhausen, Germany) at concentrations of 3, 1, and 0.3 per well and stimulated with allogeneic feeder cells and recombinant human interleukin-2 (rhIL-2; Red Swan, Utrecht, The Netherlands).…”

Dendritic Cells: Immune Saviors or Achilles' Heel?

“…Bacteria can be engineered to express gene products of interest at the cell surface, in the cytosol, or as secreted protein (51,75,95) and can also serve as carriers for introducing Ag-encoding DNA into DCs (18,20). Recombinant Streptococcus gordonii expressed on the surface the C fragment of tetanus toxin has demonstrated an extremely high capacity to deliver Ag into human monocyte-derived DCs (12).…”

Identification of three lineages of wild measles virus by nucleotide sequence analysis of N, P, M, F, and L genes in Japan

“…In developing countries, one of the major difficulties is how to immunize infants who have maternal antibody against measles. For this purpose, a synthetic peptide vaccine is thought to be a candidate, and some laboratories have attempted to identify the B or T cell recognition epitopes of MV in order to develop an immunogenic peptide vaccine for measles [Beauverger et al, 1994;Verjans et al, 1995;Hummel and Bellini, 1995;Makela et al, 1989;Obeid et al, 1995]. Taylor et al [1991] described some different lineages of wild strains of MV co-circulating at a time, and Rota et al [1992Rota et al [ , 1994b reported that significant antigenic drift was observed in the H protein of MV and that recent isolates of MV were classified into significantly different lineages from those isolated previously.…”