The human colon cancer cell line HCT does not express any detectable HLA class I antigens on the cell surface. RNA blot analyses showed that HCT cells synthesize easily detectable levels of heavy chains as well as beta 2-microglobulin (beta 2m) transcripts. Experiments of immunoprecipitation revealed the presence of intracellular HLA heavy chains and the absence of beta 2m molecules. Sequencing studies, performed on polymerase chain reaction-mediated amplification of beta 2m-specific complementary DNAs, indicated that in HCT cells both beta 2m genes are mutated. The first mutation consists of an 11-base deletion, corresponding to the first 11 base pairs of the second exon of the beta 2m gene. This mutation alters the reading frame, starting from the third amino acid residue of the mature beta 2m protein, resulting in the synthesis of a 31-amino acid peptide with no remarkable homology to any of the sequences stored in the protein database. The second mutation is a point mutation (C----A), resulting in a UAA stop codon corresponding to the 10th amino acid residue of the mature beta 2m. Therefore, it would appear that in HCT cells the beta 2m genes have undergone two different mutational changes. This is the first molecular demonstration of beta 2m mutations in a human epithelial cell line.