In vivo loss of heterozygosity in T-cells of B6C3F1 Aprt(+/-) mice

We have used B6C3F1 mice heterozygous at Aprt (adenine phosphoribosyltransf erase) as a model to study in vivo loss of heterozygosity (LOH) in normal s plenic T-lymphocytes. APRT-deficient T-cells were selected in medium contai ning 50 mu g/ml 2,6-diaminopurine (DAP), an adenine analog that is toxic on ly to cells with APRT enzyme activity. DAP-resistant (DAP(r)) T-cell varian ts were recovered at an average frequency of 3 x 10(-5) from 21 B6C3F1 Aprt (+/-) mice. Allele-specific PCR of Aprt showed that about 70% of 122 DAP(r) colonies were caused by loss of the nontargeted Aprt allele (Aprt(+)). Ana lysis of microsatellite markers along the length of chromosome 8 suggested that mitotic recombination, or chromosome loss, with or without duplication of the remaining chromosome are the predominant mechanisms resulting in lo ss of Aprt(+). DNA sequencing of Aprt RT-PCR products from the DAP(r) varia nts that retained Aprt(+) indicated that point mutation as well as other me chanisms could cause this second class of variants. The high spontaneous fr equency of in vivo Aprt LOH in mouse T-cells, mediated by LOH mechanisms th at are also known to produce human cancers, suggests that the Aprt heterozy gous mouse is a valid model for studying the diversity of mechanisms for in vivo somatic mutagenesis. (C) 2000 Wiley-Liss, Inc.