Establishment and Maintenance of DNA Methylation Patterns in Mouse Ndn: Implications for Maintenance of Imprinting in Target Genes of the Imprinting Center

Abstract

The closely linked H19 and Igf2 genes show highly similar patterns of gene expression but are reciprocally imprinted. H19 is expressed almost exclusively from the maternally inherited chromosome, while Igf2 expression is mostly from the paternal chromosome. In humans, loss of imprinting at this locus is associated with tumors and with developmental disorders. Monoallelic expression at the imprinted Igf2/H19 locus occurs by at least two distinct mechanisms: a developmentally regulated silencing of the paternal H19 promoter, and transcriptional insulation of the maternal Igf2 promoters. Both mechanisms of allele-specific silencing are ultimately dependent on a commoncis-acting element located just upstream of theH19 promoter. The coordinated expression patterns and some experimental data support the idea that positive regulatory elements are also shared by the two genes. To clarify the organization and function of positive and negative regulatory elements at theH19/Igf2 locus, we analyzed two mouse mutations. First, we generated a deletion allele to localize enhancers used in vivo for expression of both H19 and Igf2 in mesodermal tissues to sequences downstream of the H19 gene. Coincidentally, we demonstrated that some expression ofIgf2 is independent of the shared enhancer element. Second, we used this new information to further characterize an ectopicH19 differentially regulated region and the associated insulator. We demonstrated that its activity is parent-of-origin dependent. In contrast to recent results from Drosophilamodel systems; we showed that this duplication of a mammalian insulator does not interfere with its normal function. Implications of these findings for current models for monoallelic gene expression at this locus are discussed.