A novel imprinted transgene located near a repetitive element that exhibits allelic imbalance in DNA methylation during early development.

A mouse line carrying a lacZ transgene driven by the human EEF1A1/EF1 alpha promoter was established. Although the promoter is known to show ubiquitous activity, only paternal transgene alleles were expressed, resulting in a transgene imprinting. At mid-gestation, the promoter sequence was differentially methylated, hypomethylated for paternal and hypermethylated for maternal alleles. In germline, the promoter was a typical differentially methylated region. After fertilization, however, both alleles were hypermethylated. Thus, the differential methylation of the promoter required for transgene imprinting was re-established during later embryonic development independently of the germline differential methylation. Furthermore, also a retroelement promoter closely-flanking imprinted transgene and its wild type counterpart displayed similar differential methylation during early development. The retroelement promoter was methylated differentially also in germline, but in an opposite pattern to the embryonic differential methylation. These results suggest that there might be an unknown epigenetic regulation inducing transgene imprinting independently of DNA methylation in the transgene insertion site. Then, besides CpG dinucleotides, non-CpG cytosines of the retroelement promoter were highly methylated especially in the transgene-active mid-gestational embryos, suggesting that an unusual epigenetic regulation might protect the active transgene against de novo methylation occurring generally in mid-gestational embryo.