An H-2Kb negative BL6 melanoma clone (BL6-8) was transfected with plasmids containing either the class I H-2Kb or class II H-2IAk gene in combination with the neor gene. The effects of the transfected genes on the expression of the melanoma-associated antigen (MAA) recognized by the monoclonal antibodies MM2-9B6 and MM2-3C6 and the cell surface carbohydrates recognized by 15 different lectins were studied. The original H-2Kb- clone or clones transfected with neor or class II H-2IAk genes expressed high levels of MAA and very low levels of soybean agglutinin (SBA), Griffonia simplicifolia I-B4 (GSIB4), and peanut agglutinin (PNA) lectin-binding sites. In contrast, clones that expressed high levels of the transfected H-2Kb gene completely lost the expression of MAA. In addition, these clones were characterized by the appearance of high levels of expression of the sugars specifically reacting with SBA, GSIB4, and PNA lectins. When the original BL6-8 clone was transfected with the H-2Kd gene, 25 clones subsequently isolated had relatively low expression of the transfected H-2Kd gene but high expression of the endogenous H-2Kb gene accompanied by an alteration in expression of the MAA and lectin binding identical with patterns common for H-2Kb+ melanoma cells. These changes were not due to the transfection, plasmid construction, or place of insertion, since similar phenotypic characteristics were found in H-2Kb+ but not H-2Kb- clones isolated from the N-methyl-N'-nitro-N-nitrosoguanidine-treated BL6T2 or parental BL6 melanoma lines. In total, 73 BL6 melanoma clones were investigated and all of the 41 H-2Kb+ clones displayed loss of MAA and appearance of SBA, GSIB4, and PNA-binding sugars. None of the 32 H-2Kb- clones showed these changes. This study indicates that the class I H-2Kb gene product might alter several phenotypic properties of BL6 melanoma cells. The mechanisms of these changes remain unknown. We consider that these effects of the class I H-2Kb gene are indirect, involving interactions with the B-tropic ecotropic retrovirus specific for melanomas of C57BL/6 mice origin.