Early in development, a highly coordinated maternal-to-zygotic transition ?reprograms? the embryo to a pluripotent cellular identity, capable of giving rise to all subsequent cell types in the developing organism. How cells are induced to pluripotency is still poorly understood, and yet has enormous implications, not only for understanding fundamental developmental and gene regulatory processes, but also for disease modeling and therapies. The long-term goal is to decipher mechanisms and principles conserved across vertebrates that guide pluripotency induction. To begin to address this question, this proposal aims to elucidate how pluripotency factors provided to the egg activate the first genes from the embryonic genome in the model vertebrate zebrafish. Zebrafish produce large clutches of externally developing embryos and express many key regulators of pluripotency that are conserved with humans, making them an ideal experimental system to discover novel regulatory paradigms that induce vertebrate pluripotency. This proposal has three primary goals. First, the regulatory logic underlying the specificity of embryonic gene activation will be deciphered. Maternal pluripotency factors act on a transcriptionally quiescent embryonic genome to initially activate only a few hundred genes, prior to the activation of thousands of genes hours later. Cutting-edge techniques will be used to map pluripotency factor binding genome wide, at earlier developmental stages than have previously been profiled. Binding patterns will be compared in the vicinity of early- versus later activated genes, and distinguishing properties of their respective gene regulatory sequences will be extracted. Second, novel regulators of embryonic genome activation will be deduced by integrating multiple sources of genomics data. These factors will be evaluated using innovative loss-of-function strategies to measure their contributions to embryonic reprogramming and pluripotency. Third, this proposal will uncover post-transcriptional mechanisms that regulate the maternal pluripotency factors themselves. Biochemical and genetic approaches will be used to discover factors that bind maternal mRNA to positively or negatively affect their expression, and in turn exert precise temporal control over pluripotency induction. Together, these analyses will further elucidate the gene regulatory logic of the early vertebrate embryo and provide a deeper understanding of how pluripotent identities are induced.
This proposal aims to uncover regulatory paradigms underlying how reprogramming factors trigger widespread gene expression changes, by deciphering the regulatory logic of the early embryo that leads to genome activation and the establishment of pluripotent stem cells. Transcriptional reprogramming underlies disorders of cell identity such as cancer, and understanding how gene expression can be reprogrammed is fundamental to developing strategies to treat such disorders. Additionally, the potential to precisely manipulate cellular identity through transcriptional reprogramming has enormous implications for disease modeling and therapies, thus motivating the need to discover novel vertebrate mechanisms that contribute to this process.
