A subset of genes in mammals is regulated by genomic imprinting, a process that results in unequal expression of the maternal and paternal alleles of this class of genes. Imprinted genes are hypothesized to expiain why nuclear contributions from both parents are required for normal mammalian development. Furthermore, imprinting plays a role in the transmission of a number of human genetic diseases, including Beckwith-Wiedemann Syndrome (BWS), Silver-Russell Syndrome (SRS), Prader-Willi and Angelman Syndrome, in that the sex ofthe parent that transmits the affected gene(s) determines whether offspring will develop the disease. Aben-ant imprinted gene expression is also involved in the establishment or progression of cancers, inciuding Wilms tumors. The objective of this proposal is to investigate the- mechanism by which parental identity of imprinted genes is established and maintained. The studies will employ the conserved H19/lgf2 locus. The imprinting of H19, which produces a non-coding RNA from the maternally-derived allele, and the linked and oppositely imprinted growth-promoting Igf2 gene is mediated through the 2 kb imprinting control region (ICR) and shared enhancers. The ICR acts as a methylation- senstitive, CTCF-dependent insulator. When unmethylated on the maternal allele, the insulator allows H19 exclusive access to the enhancers. In contrast, a methylated paternal insulator enables Igf2 to engage the enhancers. This proposal will investigate the mechanism of H19/lgf2 imprinting through the following experiments. Individuals with BWS, sporadic Wilms tumors and SRS have been identified that have microdeletions and epimutations in the human ICR and aberrant imprinted regulation of H19 and Igf2. We wiil generate mouse models and human IPS cells with these mutations and study the mechanism of loss of imprinting. We will also investigate the function of a conserved non-coding RNA located between H19 and Igf2 and the H19 micro RNA, miR-675, using mutations constructed at the mouse locus. We will additionally explore the role of TET1 in the establishment and erasure of ICR methylation. Last, the imprinting mechanism will be studied at the Grb10 locus, which may employ aspects of H19 imprinted regulation.
(See insfruc:tions): Imprinted genes are critical for normal mammalian development, behavior and energy homeostasis. This genes have genetic or epigenetic mutations in a number of human syndromes and cancer. The experiments in this proposal will model such newly identified mutations in individuals with Beckwith-Wiedemann Syndrome and Silver-Russell Syndrome, providing a better understanding ofthe etiology ofthe disease. PROJECT/PERFORIVIANCE SITE(S) (if additional space Is needed, use
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