Most genes in mammals are expressed equally from the maternally and paternally inherited alleles. However, some genes are imprinted, and are expressed from only one parental allele. One consequence of imprinting is that a single mutation or deletion in the expressed allele, will result in the absence of a gene product despite the presence of a wild- type, but silent allele. This is the type of mutation involved in the genetic diseases Prader-Willi Syndrome (PWS) and Angelman Syndrome (AS), which represent distinct clinical phenotypes resulting from opposite patterns of genomic imprinting of human chromosome 15q11-q13. The experiments described in this proposal will firmly establish the mouse as a model system for imprinting of human 15q11-q13 and begin to address the more intricate mechanistic questions that may be unapproachable in humans. The first specific aim is to create mouse models of human PWS and AS imprinting mutations to test the hypothesis that the region upstream of the Snrpn gene serves as a critical imprinting center, responsible for the global regulation of multiple imprinted genes. If we find evidence that there is an imprinting center, we will take advantage of our ability to create independent but identical deletion mutations on both parental alleles in mice to investigate the mechanism of the imprinting center as well as the relevance of the non-overlapping AS and PWS deletions. The second specific aim is to determine if either of the two Snrpn open reading frames play a role in the imprinting process and evaluate each open reading frame's contribution to PWS. The third specific aim will focus on the imprinting machinery in and around the Snrpn locus as a means of determining how parental identity is assigned. This will involve determining if there is parent-specific methylation differences and if there are any nearby transcriptional units. The fourth specific aim is to develop a functional assay to identify the cis-acting elements that cause the Snrpn gene to be imprinted and subsequently determine if the minimal identified sequence are sufficient to confer paternal-specific expression to another gene.
