Assessment of major histocompatibility complex class I interaction with Epstein‐Barr virus and human immunodeficiency virus peptides by elevation of membrane H‐2 and HLA in peptide loading‐deficient cells

Abstract

Earlier findings indicate that peptides can affect the expression of major histocompatibility complex (MHC) class I molecules on the surface of cells with defective peptide loading mechanism. We have used peptide induced increase of class I antigen expression to assess peptide interaction with MHC class I molecules. A panel of 41 overlapping synthetic peptides derived from the human immunodeficiency virus‐1 (HIV‐1) gag protein and 33 nonoverlapping peptides from Epstein‐Barr virus (EBV) proteins EBNA‐1, 2, 3, 4, 5, 6, LMP, BZLF2, BILF2, BSLF2, BALF4 and BcLF1 was assessed for the ability to enhance the expression of HLA‐A2.1, H‐2D_b, K_b and D_d on the murine RMA‐S and human 721.174/T2 (.174/T2) lines by indirect immunofluorescence. Considering doubling of the fluorescence intensity in the peptide‐treated samples as positivity, 6 of 39 HIV and 1 of 32 EBV peptides were found to bind to A2.1, 6 of 39 HIV gag and 7 of 16 EBV peptides to D_b, 8 of 39 HIV gag and 5 of 16 EBV peptides to K_b and 2 of 39 HIV gag and 1 of 17 EBV peptides to D_d. The sensitivity of the method is comparable to the in vitro class I assembly assay with conformation‐dependent monoclonal antibody and is more discriminating than the solid‐phase assay. Due to its simplicity this method can also serve for testing large peptide panels for binding capacity to various class I molecules. Moreover, the method provides information about the relevance of in vitro tests for class I assembly in living cells.