Loss of heterozygosity (LOH) plays an important role in the expression of recessive mutations in mammalian cells. To gain insight into the rate and mechanisms of LOH the autosomal HLA-A gene was used as a model system. Spontaneous HLA-A2 mutants originated with a rate of respectively 4.1 x 10(-6) and 6.9 x 10(-6) per cell per generation in TK6 and WI-L2-NS, two isogenic lymphoblastoid cell lines which differ in TP53 status. The rate of loss of HLA-A2 is 10-50 times higher compared to the mutation rate of the X-linked HPRT gene. The homozygous TP53 mutation in WI-L2-NS had no effect on the rate of HLA-A2 loss or the spectrum of these mutations. Microsatellite analysis of most of the HLA-A2 mutants (84%) showed LOH for multiple markers on chromosome arm 6p telomeric of a recombination breakpoint, LOH for all 6p markers, or LOH for markers on both the 6p- and 6q-arms. Cytogenetic analysis showed that these mechanisms gave mutant cells which harbored two intact chromosomes 6 and which were indistinguishable from non-mutant cells. Therefore, loss of HLA-A2 is mainly caused by somatic recombination (33-50%) or chromosome loss with duplication of the remaining chromosome (34-40%). These findings correspond to the mechanisms behind loss of the wild-type RBI allele in retinoblastoma and suggest that both somatic recombination and chromosome loss followed by duplication contribute to tumorigenesis.