CHARACTERIZATION OF AN EPITOPE OF THE HUMAN CYTOMEGALOVIRUS PROTEIN IE1 RECOGNIZED BY A CD4(-CELL CLONE() T)

CD4(+) T cells specific for human cytomegalovirus (HCMV) IE1 protein a re potential effecters of the control of HCMV infection through cytoki ne production. Better knowledge of major histocompatibility complex (M HC)-peptide-T cell receptor (TcR) interactions in the CD4(+) T cell re sponse should result in a better design of immunizing peptides and is a prerequisite for the development of vaccines or anti-cytomegalovirus therapy. In this study, the recombinant protein comprising residues 8 6-491 encoded by exon 4 of IE1 (GST-e4) was cleaved by enzymatic diges tion and analyzed by high pressure liquid chromatography-mass spectros copy (HPLC-MS). We identified the 14-residue epitope 162-DKREMW/MACIKE LH-175 recognized by an HLA-DRS-restricted clone, BeA3. Synthetic elon gated, truncated and di-Ala-substituted peptides of the 18-mer IE1 158 -IVPED KREMWMACIKE LH-175 sequence were used to analyze the amino acid motifs involved in binding to HLA-DRS and recognition by the BeA3 clo ne. Substitutions which abolished (MW --> AA), or decreased (RE --> AA and MA --> AA) T cell clone proliferation, cytokine production and cy totoxicity were identified. Loss of T cell function induced by the MW --> AA substitution was associated with poor HLA-DRS binding. Decrease d T cell function (RE --> AA and MA --> AA) was associated with good H LA-DRS binding, which suggested that these motifs were involved in TcR binding. Other substitutions induced potentiation of the T cell done response: the IV --> AA substitution induced stronger proliferation, b ut equivalent cytokine production, when compared with the reference pe ptide IE1 (158-175). CI --> AA substitution induced strong potentiatio n of HLA-DR8 binding, proliferation and interferon-gamma and interleuk in4 production, possibly due to the removal of negative effects of Cys , Ile, or both side chains. Cytotoxicity was not improved by any subst itutions. Our results show modulation of the CD4(+) T cell response ac cording to the peptide residues involved in the HLA-DR8-peptide-TcR in teraction.