Methylation of adenovirus genes in transformed cells and in vitro: influence on the regulation of gene expression?

An inverse correlation has been described between the levels of DNA methylation in specific segments of adenovirus DNA integrated into the genomes of transformed and tumor cells and the extent to which these segments are expressed as messenger RNA. In the adenovirus type 2 (Ad2)-transformed hamster cell lines HE2 and HE3, the virus-specific DNA binding protein (DBP) is not expressed, and the DNA in the DBP gene is completely methylated in all 5'-CCGG-3' sites. At least part of the late promoter/leader sequence of the DBP gene is present in cell lines HE2 and HE3. In line HE1, on the other hand, the DBP is expressed, and the DNA in the DBP gene is unmethylated at the 5'-CCGG-3' (HpaII) sites. The late promotor/leader sequence of the DBP gene is expressed in cytoplasmic RNA isolated from line HE1. The effect of DNA methylation has also been tested in vitro in a microinjection system using Xenopus laevis oocytes. Unmethylated DNA fragments of Ad2 (E2a region) have been found to serve as active templates. When the same fragments are methylated at the 5'-CCGG-3' sites by the HpaII DNA-methyltransferase, viral RNA synthesis is inhibited upon microinjection into oocyte nuclei. These results provide direct evidence for the notion that DNA methylated at highly specific sites is somehow involved in the regulation of gene expression.