Parental allele-specific chromatin configuration in a boundary-imprinting-control element upstream of the mouse H19 gene

The mouse H19 gene is expressed from the maternal chromosome exclusively. A 2-kb region at 2 to 4 kb upstream of H19 is paternally methylated througho ut development, and these sequences are necessary for the imprinted express ion of both H19 and the 5'-neighboring Igf2 gene. In particular, on the mat ernal chromosome this element appears to insulate the Igf2 gene from enhanc ers located downstream of H19, We analyzed the chromatin organization of th is element by assaying its sensitivity to nucleases in nuclei. Six DNase I hypersensitive sites (HS sites) were detected on the unmethylated maternal chromosome exclusively; the two most prominent of which mapped 2.25 and 2.7 5 kb 5' to the H19 transcription initiation site. Five of the maternal HS s ites were present in expressing and nonexpressing tissues acid in embryonic stem (ES) cells. They seem, therefore, to reflect the maternal origin of t he chromosome rather than the expression of H19, A sixth maternal HS site, at 3.45 kb upstream of H19, was detected in ES cells only. The nucleosomal organization of this element was analyzed in tissues and ES cells by microc occal nuclease digestion. Specifically on the maternal chromosome, an unusu al and strong banding pattern was obtained, suggestive of a nonnucleosomal organization. From our studies, it appears that the unusual chromatin organ ization with the presence of HS sites (maternal chromosome) and DNA methyla tion (paternal chromosome) in this element are mutually exclusive and refle ct alternate epigenetic states. In addition, our data suggest that nonhisto ne proteins are associated with the maternal chromosome and that these migh t be involved in its boundary function.