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.
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-2Db, Kb and Dd 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 Db, 8 of 39 HIV gag and 5 of 16 EBV peptides to Kb and 2 of 39 HIV gag and 1 of 17 EBV peptides to Dd. 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.
Stuber
G
G
Department of Tumor Biology, Karolinska Institute, Stockholm, Sweden.
Modrow
S
S
Höglund
P
P
Franksson
L
L
Elvin
J
J
Wolf
H
H
Kärre
K
K
Klein
G
G
D000595:Amino Acid Sequence; D000818:Animals; D000956:Antigens, Viral / Q000276:immunology*; D004268:DNA-Binding Proteins / Q000276:immunology*; D019309:Epstein-Barr Virus Nuclear Antigens; D015683:Gene Products, gag / Q000276:immunology*; D006183:H-2 Antigens / Q000032:analysis*; D015497:HIV-1 / Q000276:immunology*; D015789:HLA-A2 Antigen / Q000032:analysis*; D004854:Herpesvirus 4, Human / Q000276:immunology*; D006801:Humans; D051379:Mice; D008810:Mice, Inbred C57BL; D008969:Molecular Sequence Data; D014764:Viral Proteins / Q000276:immunology*
D000956:Antigens, Viral; D004268:DNA-Binding Proteins; D019309:Epstein-Barr Virus Nuclear Antigens; D015683:Gene Products, gag; D006183:H-2 Antigens; D015789:HLA-A2 Antigen; D014764:Viral Proteins