Loss of critical palindromic transgene promoter sequence in chemically induced Tg.AC mouse skin papillomas expressing transgene-derived mRNA

The Tg.AC transgenic mouse carries a v-Ha-ras transgene. Skin papillomas develop in Tg.AC mice upon repeated dermal application of tumor promoters and carcinogens. The transgene is inserted at a single site on chromosome 11 in a multiple-copy array. Although most of the greater than or equal to 40 copies are arranged in a direct-repeat orientation, two copies of the transgene are inserted in a palindromic, inverted-repeat orientation. Deletion of the palindromic transgene promoter sequence is associated strongly with and diagnostic of loss of phenotypic responsiveness to Tg.AC papillomagens, such as 12-O-tetradecanoylphorbol-13-acetate (TPA). Unexpectedly, a loss of palindromic transgene sequence, in the absence of an observable reduction in copy number of the direct-repeat-oriented transgene sequence, is seen in DNA from papillomas when compared to genomic DNA from tail clips or skin samples away from the application site. Transgene-derived transcripts were detectable in all Tg.AC papillomas sampled. The transgene locus was hypomethylated in papillomas but not in samples from tail clips from the same animal or from skin samples away from the application site in responder Tg.AC mice, as shown by loss of resistance to digestion by Hpall. A cell line derived from a Tg,AC squamous cell carcinoma showed complete loss of the palindromic transgene sequence, hypomethylation of the transgene locus, and strong expression of v-Ha-ras mRNA. These data indicate that the palindromic transgene sequence, which appears to be necessary for initial responsiveness to tumorigens, may be susceptible to deletion during rapid cellular proliferation and is not required for transgene expression in later phases of papilloma growth. Published 2001 Wiley-Liss, Inc.