Characterization of the specificity and genetic restriction of human CD4+ cytotoxic T cell clones reactive to capsid antigen of rubella virus

Abstract

Using 11 overlapping synthetic peptides covering more than 95% of the amino acid sequence of capsid protein of rubella virus, 7 CD4+ T cell clones (R10, R11, R18, A2, A10, A11, and A12) isolated from 2 rubella seropositive donors reacted strongly to rubella capsid peptides C6 (residues 119-152), C9 (residues 205-233), or C11 (residues 255-280), respectively, in both proliferation and cytotoxicity assay. Truncated peptides C6E (residues 125-139), C9B (residues 205-216), and C11E (residues 260-272) were shown to be involved directly to the T cell determinants of C6, C9, and C11, respectively. Genetic restriction of these T cell clones was analyzed by using human cell lines with various HLA-DR phenotypes as targets and/or antigen-presenting cells in cytotoxicity assay and/or proliferation assays. The results indicated that the recognition of peptide C6 by T cell clones (R11 and R18) was associated with DRw9 molecule, while the HLA restriction element of the responses of other T cell clones (A2 and A11, A10, and A12) that reacted with peptide C9 or C11 was DR4 molecule. However, there may be a cross-recognition by the T cell clone (A12) between DR1 and DR4 subtypes.

Analysis of overlapping T- and B-Cell antigenic sites on rubella virus E1 envelope protein influence of HLA-DR4 polymorphism on T-cell clonal recognition

“…The T-cell clones (ATRVC2 and AT177C5) used for this study were isolated from two CD4+CD8 T-cell lines derived from a single RVseropositive donor as described previously [22,24,25]. Briefly, PBMNCs were incubated with UV-inactivated RV (5 × 105 FFU/ml) or the SP E1(273-291), which was used at a final concentration of 15 ~g/ml in complete RPMI-1640 medium containing 2 mM L-glutamine, 25 mM N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid (Hepes), 50 mM penicillin, 50 mM streptomycin, and 5 × 10 -5 M 2-mercaptoethanol, which was also supplemented with 10% autologous plasma.…”

“…HCLs, EBV-BLs BSM, YAR, JHa, PF, KB, HOM2, MAT, CRI, Ig-38, MST, SST, and KT2, which are homozygous at the HLA region, with different HLA-DR phenotypes or DR4 subtypes (Tables 4 and 5 Cell-mediated cytotoxicity assay. A standard 51Cr-release assay was used to measure CTL responses [22,23,25]. For use as CTL targets, EBV-BL or HCL cells (1 × 106) were sensitized by overnight incubation at 37°C with 1 × 106 FFU of UV-inactivated RV, or 5 I-~g/ml rE 1BV, or 10 gtM of SP in 1 ml of complete RPMI medium.…”

Cross-reactive rubella virus and glutamic acid decarboxylase (65 and 67) protein determinants recognised by T cells of patients with Type I diabetes mellitus

“…To examine the crossrecognition of RV antigens and beta-cell antigens by T-cells from different patient cohorts, a panel of 15 peptides (V1-V15) of RV structural protein E1, E2, and C were selected for this study (Table 2). Peptides V1, V4, V5, V8, V11, V13, V14, V15 are antigenic peptides containing epitopes identified by T-cells from RV seropositive healthy donors or RV vaccinees or both in our previous studies [32,33,43,44]. RV peptides V2, V3, V6, V7, V9, V10, V12 were shown earlier to be recognised by peripheral blood T-cells of CRS patients [31] and included five (V3, V6, V7, V9, V12) which stimulated cellular responses of a CRS patient who also had Type I diabetes.…”

Point Mutation of a Rubella Virus E1 Protein T-Cell Epitope by Substitution of Single Amino Acid Reversed the Restrictive HLA-DR Polymorphism: A Possible Mechanism Maintaining HLA Polymorphism

“…Two kinds of EBV-BL were used as targets in 51Cr release assay. EBV-BLs AT (autologous to T-cell clone AT177C5) was established in our laboratory by the method described previously (22)(23)(24). HCLs, EBV-BLs BSM, YAR, SST, JHa, MT, KT2, and KT3, which are homozygous at the HLA region, and EBV-BL P9B, which is heterozygous at HLA-DR (DR4/DR2), with different HLA-DR phenotypes or DR4 subtypes (Table 2) were provided kindly by Dr. G.T.…”

“…A standard 51Cr release assay was used to measure CTL responses (22)(23)(24). Briefly, for use as CTL targets, EBV-BL or HCL cells (1 X 106) were incubated at 37°C with 10 pM.…”