? Defects in parent of origin dependent, or """"""""imprinted"""""""", gene expression are associated with a number of human diseases including Beckwith-Wiedemann Syndrome (BWS), Prader-Willi Syndrome, Angelman Syndrome and several types of cancer. The purpose of this study is to examine the genetic and epigenetic mechanisms underlying loss of imprinting (LOI) at H19 and IGF2 that has been observed in patients with BWS. LOI in these patients was attributed to microdeletions in the imprinting control region (ICR) of H19 and Igf2. Previous studies have shown that binding sites for the insulator protein CTCF present at the ICR are necessary for imprinting control. Furthermore, the formation of higher-order chromatin at the H19/lgf2 locus has been implicated in imprinting control. This work will address the role of the spacing and pattern of CTCF sites in imprinting. To achieve the proposed aims, mice will be generated carrying the following microdeletions at the H19/lgf2 ICR: (i) 0.8 kb between CTCF sites 2 and 3; and 1.3 kb between sites 1 and 4. The effects of these mutations on imprinting at H19 and Igf2 will be determined using allele-specific assays to test expression levels of H19 and Igf2 and methylation at the ICR and H19 promoter. It will also be determined whether these mutations interfere with CTCF binding or posttranslational histone modifications by allele-specific chromatin immuno-precipitation assays. Whether the mutations affect the formation of a higher-order chromatin structure at the H19/lgf2 locus will be tested by chromosome conformation capture (3C) assays. As the number and spacing of CTCF sites present at the ICR differs between mice and humans, it is necessary to generate a model to directly test the effect of mutations observed in patients. Therefore, mice that carry the human ICR (hlC1) in place of the mouse ICR will be generated and tested for whether hlC1 can regulate imprinting of mouse H19 and Igf2. Finally, a recent study revealed that the H19/lgf2 ICR is transcribed. Here, the level and pattern of ICR transcription will be determined and how it relates to the level and pattern of imprinted expression at H19 and Igf2. Moreover, the role of CTCF in ICR transcription will be investigated by studying ICR transcription in conditional CTCF knockout oocytes. Research pertaining to the association between epigenetics and human disease is a relatively new and fast growing field. An increasing number of diseases are being attributed to epigenetic defects, however, the underlying mechanism is still poorly understood. The research proposed here will open new doors to our understanding of the genetic and epigenetic mechanisms underlying genomic imprinting and thereby have significant impact on the study and treatment of the associated human diseases. ? ? ?
| Freschi, Andrea; Hur, Stella K; Valente, Federica Maria et al. (2018) Tissue-specific and mosaic imprinting defects underlie opposite congenital growth disorders in mice. PLoS Genet 14:e1007243 |
| Hur, Stella K; Freschi, Andrea; Ideraabdullah, Folami et al. (2016) Humanized H19/Igf2 locus reveals diverged imprinting mechanism between mouse and human and reflects Silver-Russell syndrome phenotypes. Proc Natl Acad Sci U S A 113:10938-43 |
| Ideraabdullah, Folami Y; Thorvaldsen, Joanne L; Myers, Jennifer A et al. (2014) Tissue-specific insulator function at H19/Igf2 revealed by deletions at the imprinting control region. Hum Mol Genet 23:6246-59 |
| Ideraabdullah, Folami Y; Bartolomei, Marisa S (2011) ZFP57: KAPturing DNA methylation at imprinted loci. Mol Cell 44:341-2 |
| Ideraabdullah, Folami Y; Abramowitz, Lara K; Thorvaldsen, Joanne L et al. (2011) Novel cis-regulatory function in ICR-mediated imprinted repression of H19. Dev Biol 355:349-57 |
