MOUSE MODEL FOR HUMAN LOSS OF HETEROZYGOSITY

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Loss of heterozygosity (LOH) is well documented in cancers. When LOH occurs at tumor suppressor loci, a recessive (mutant) allele becomes functionally homozygous or hemizygous, leading to a relative loss of cellular growth regulation and, ultimately, tumorigenesis. Environment genotoxicants may induce LOH leading to a large fraction of cancers and inherited disease. Thus, it is important to understand mechanisms of LOH and to evaluate environments and agents in terms of their ability to induce LOH in whole animals. Unfortunately, existing whole animal (e.g., transgenic mice) assays do not detect several of the classes of events that produce LOH. Thus, we require practical and sensitive assays that actually detect and quantitate LOH PER SE, independent of mechanism. Using targeted homologous recombination of ES cells, we produced mice that are heterozygous at the adenine phosphoribosyltransferase (Aprt ) locus. In vivo LOH at this locus produces cellular loss of APRT activity, which yields a selectable phenotype when tissues are dissociated and cells cultured. Thus, we have an assay that measures in vivo LOH at an endogenous locus in normal animals. Further, the analysis of polymorphic loci flanking Aprt, Dna sequencing of mutant Aprt genes, and analysis of gene organization and chromosome structure with fluorescent in situ hybridization enables the determination of molecular mechanisms producing LOH. We will examine the spontaneous rate and mechanism of LOH in various mouse tissues, in mice with different genotypes which may predispose cells to LOH and tumorigenesis, and after exposure to genotoxic agents.