Molecular Mechanisms Employed by TRIM28 to Maintain Epigenetic Information through Different Stages of Embryonic Development

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How information in genomes is decoded during embryogenesis is one of the greatest mysteries of humankind. Central to this problem is an understanding of how gene expression is regulated. Certain genes are controlled by chemical modifications that are introduced before fertilization. These marks to DNA (also called chromatin marks or genomic imprints) in eggs and sperm are transmitted from parents to children and constitute an important mechanism for the control of gene expression across generations. While research in the last 50 years has uncovered the complexity of chromatin marks, information is still lacking about the factors that recognize and interpret these marks to activate or repress the imprinted genes. Preliminary evidence has revealed a novel role for a specific protein as a regulator of imprinted gene expression. By further defining the requirements for involvement of this protein, this research program will expand fundamental knowledge of the processes and factors that mediate control of gene expression across generations. In addition to enhancing understanding of a fundamental problem in embryonic development, this program of study fosters research by students of all levels. Additionally, both the lead researcher and students participate in several outreach activities to disseminate knowledge of developmental biology. Shortly after fertilization, mammalian embryos undergo a genome-wide demethylation event that involves the enzymatic removal of methyl groups from methyl-cytosine DNA residues. The removal of these epigenetic marks allows the reprogramming of the zygotic genome, an essential process for embryogenesis to begin. However, a number of genomic loci, including imprinted genes, need to preserve epigenetic information for the new organism to develop properly. The transcriptional repressor TRIM28 has recently emerged as a key regulator of imprinted gene expression. However, the mechanisms employed by this transcription factor are still unclear. This proposal aims to resolve the requirements of TRIM28 for genomic imprinting at different developmental stages and the molecular mechanisms used by this transcriptional regulator to maintain epigenetic information in developing mouse embryos. Experiments in Specific Aim 1 will analyze the stage-specific requirements of TRIM28 for imprinting control by analyzing the effects of conditional depletion of Trim28 at different times during embryonic development. In Specific Aim 2, the mechanisms by which TRIM28 controls secondary DMR methylation at the Dlk1-Gtl2 locus will be investigated through epistatic analysis. Results from these experiments will provide novel insight not only into the roles of TRIM28, but also into the mechanisms that regulate genomic imprinting.