The frequent loss of beta2-microglobulin (beta2-mu) in malignant cells has stimulated interest in the functional characteristics of beta2-mu-free HLA class I heavy chains, since this information contributes to assess the impact of beta2-mu abnormalities on the interaction of malignant cells with immune cells. Therefore, the present study has investigated the ability of beta2-mu-free HLA class I heavy chains to modulate NK cell-mediated lysis of melanoma cells and to present melanoma-associated antigen (MAA)-derived peptides to HLA class I-restricted, MAA-specific cytotoxic T lymphocytes (CTL). Beta2-mu-free HLA class I heavy chains were induced on beta2m null FO-1 cells by sequential incubation with IFN-alpha for 48 h at 37 degrees C and for 24 h at 26 degrees C. Transfection of cells with a wild-type H-2Ld gene (FO-1Ld) enhanced the induction of beta2-mu-free HLA class I heavy chains under such experimental conditions. Beta2-mu-free HLA class I heavy chains expressed on the cell membrane did not protect the B2m null FO-1 cells from NK cell-mediated lysis. Furthermore, FO-1 cells which express beta2-mu-free HLA-A2 heavy chains following transfection with a wild-type HLA-A2 gene were not lysed by HLA-A2-restricted, MAA-specific CTL lines and clones. These results indicate that association with beta2-mu is required for interaction of HLA class I molecules with NK inhibitory receptors and for peptide presentation to CTL.