Imprinting of Igf2 and H19 from a 130KB YAC Transgene

Justin Ainscough
Wellcome/CRC Institute

To investigate the mechanism(s) of genomic imprinting in mice, we are studying two closely linked, oppositely imprinted genes, insulin-like growth factor II (Igf2) and H19, located on distal chromosome 7. Although Igf2 is expressed from the paternal allele, and H19 is maternally expressed, they show very similar spatial and temporal patterns of expression which suggests that they may share common regulatory elements. In a previous study we performed an extensive characterization. of the 130 kb genomic domain around Igf2 and H19, in which we identified a strongly hypersensitive region located centrally between the genes which is a candidate for an imprinting control element.

As an initial step towards investigating the regulatory elements responsible for controlling imprinted expression in this domain, we generated transgenic mouse lines with a 130 kb Igf2/H19 YAC. We found that both genes were appropriately imprinted and were expressed in a wide range of tissues of both mesodermal and endodermal origin. Therefore, many of the elements responsible for enhanced expression and imprinting of these two genes are located within this 130 kb.

To further investigate the function of the intergenic "element" we have flanked the hypersensitive sites on the YAC with loxP recombination sites which has enabled us to specifically delete this region after integration into the mouse genome. This allows a direct comparison of transgene activity, with and without the element, at the same location in the mouse genome. The analysis has revealed a tissue specific silencer function for this element, where the Igf2 gene is reactivated from the maternal allele in skeletal muscle. No difference is seen in the reactivation pattern whether the region is deleted prior to germline transmission or after fertilization, demonstrating that the element is essential for maintaining the silent state of the maternal Igf2 allele.