Role of Gadd45 in pluripotency and murine embryogenesis

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It is well established that reprogramming of somatic cell nuclei during animal cloning requires extensive DNA demethylation of pluripotency genes such as nanog or oct4, which are silenced in somatic cells by DNA methylation. However, little was known about the molecular mechanisms underlying DNA demethylation. We recently found that Gadd45a is a key regulator of active DNA demethylation of oct4 in Xenopus oocytes and that it acts by promoting DNA repair. Here we propose to study the biological role of Gadd45-mediated DNA demethylation in murine embryonic stem (ES) cells and mouse embryos. While individual Gadd45a, -b and -g knock out mice have been generated previously, these mice are all viable and fertile, likely due to functional redundancy. To circumvent redundancy of Gadd45 genes, we have developed a dominant negative (dn) Gadd45. Using this tool, we will study the role of Gadd45 in pluripotency by overexpressing dnGadd45 (loss of function) as well as wild type Gadd45 (gain of function) in ES cells. In parallel, we will generate and characterize transgenic mice using dnGadd45 as well as wild type Gadd45 and revisit the role of this gene in early mouse embryogenesis.