Influence of MHC class 1:peptide interaction on antigen presentation in normal and malignant cells

Abstract: INFLUENCE OF MHC CLASS l:PEPTIDE INTERACTION ON ANTIGEN PRESENTATION IN NORMAL AND MALIGNANT CELLS Qian-Jin Zhang Doctoral dissertation from the Microbiology and Tumor Biology Center, Karolinska Institute, Stockholm, SwedenThe presentation of MHC class I:peptide complexes in normal andmalignant cells was studied. An HLA A11 binding motif was defined bycomparing synthetic peptide analogues of a known cytotoxic T Iymphocyte(CTL) target epitope for induction of surface A11 expression and triggering ofcytotoxic activity. The motif predicts the presence of hydrophobic aminoacids in position 2 (P2), small amino acids in P3 and P6 and Lys in P9. Theoff-rates of P2 analogues discriminated between "apparent" and "true"binders that formed complexes with a half life longer than 72 hours. Thepersistence of MHC:peptide complexes the surface of antigen presenting cellswas shown to correlate with the immunogenicity two A11-restrictedepitopes of the Epstein-Barr virus (EBV) nuclear antigen (EBNA)4(designated IVT and AVF, respectively). Molecular modeling and alaninescanning mutagenesis of the IVT and AVF peptides demonstrated thatsolvent exposed peptide side chains affect CTL recognition as well asantibody binding suggesting that allospecific antibodies recognize thecomplexes in a fashion similar to T cell receptors. Class I molecules areimportant for target recognition by T cells as well as natural killer (NK) cells.Overexpression of the c-myc oncogene increased the sensitivity of EBVtransformed Iymphoblastoid cell lines (LCLs) to NK lysis. The reactivity withallospecific antibodies was significantly reduced suggesting that a differentset of antigenic peptides may occupy the class I groove, resulting in inabilityto deliver a negative regulatory signal to NK cells. Defective presentation ofendogenously expressed EBNA4 was demonstrated in A11 positive Burkitt'sIymphoma (BL) lines. The defect was not overcome by cytosolic expressionof the preformed epitope. The tumor cell expressed low levels of thetransporter associated with antigen processing (TAP) and behaved poorly ina streptolysin-O mediated peptide translocation assay but presentation wasnot restored by upregulation TAP activity following treatment with IFN-y.Efficient maturation of class I molecules to Endo H resistant species wasdemonstrated in pulse-chase experiments. The results localize the defect tofunctions downstream of the generation of antigenic peptides but upstreamof TAP-dependent peptide transport and class I assembly and maturation.Key words: Cytotoxic T lymphocytes, Natural Killer cells, Epstein-Barr virus,MHC class I, class I binding motifs, T cell receptor, antigen presentation,immunodominance .ISBN 91-628-2100-8