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.