beta 2-Microglobulin (beta 2-mu) gene-null human melanoma FO-1 cells display lower reactivity with anti-HLA class I monoclonal antibodies (mAb) following transfection with a wild-type mouse beta 2-mu gene (referred to as FO-1C cells) than following transfection with a wild-type human beta 2-mu gene (referred to as FO-1H cells). Furthermore, binding assays with a panel of anti-HLA class I mAb detected higher reactivity of FO-1C cells with mAb TP25.99 than with mAb CR1-S63, CR10-215, CR11-115, TP67, and W6/32 but similar reactivity of FO-1H cells with all the mAb tested. While mAb TP25.99 recognizes a determinant expressed on beta 2-mu-free and beta 2-mu-associated HLA class I heavy chains, the remaining mAb recognize determinants expressed only on beta 2-mu-associated HLA class I heavy chains. The differential effects of mouse and human beta 2-mu on the reactivity with anti-HLA class I mAb of FO-1 cells reflect more than one mechanism. Besides abnormalities in the processing of HLA class I heavy chains associated with mouse beta 2-mu, this molecular complex appears to be unstable on the plasma membrane of FO-1 cells. To analyze the interaction of mouse beta 2-mu with HLA-A and -B antigens, the HLA phenotype of FO-1 cells was determined, using a combination of isoelectric focusing analysis of antigens immunoprecipitated from radiolabeled cells with mAb to monomorphic determinants of HLA class I antigens, binding assays with a limited number of mAb recognizing HLA class I allospecificities, and sequence-specific oligonucleotide probe typing. Although association with mouse beta 2-mu does not cause marked differences in the expression of HLA-A25 and -B8 antigens on the cell surface of FO-1 cells, it causes a selective reduction in the expression of determinants recognized by anti-HLA-A mAb F4/72 and VF19-LL67 and by anti-HLA class I mAb W6/32 on HLA-A25 allospecificities. The differential effect of the association with mouse beta 2-mu on the antigenic profile of HLA-A25 and -B8 antigens may reflect the different characteristics of the amino acids at residue 12, which interact with residue 33 of beta 2-mu. The latter residue is the only one to differ between human and mouse beta 2-mu in the stretch of amino acids interacting with the alpha 1 and alpha 2 domains of HLA class I heavy chains.(ABSTRACT TRUNCATED AT 400 WORDS)