Mechanism for the reprogramming of gene expression in the oocytes and preimplantation embryos

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This research project consists of following 3 steps.1. Profiling of the genes which are expressed at the onset of zygotic gene activation (ZGA) at 1-cell stage.2. Analysis for the regulation of gene expression in the growing oocytes and early preimplantation embryos by examining transcription factors and epigenetic modifications.3. Functional analysis by RNAiThe results of these studies are described below1. The nascent mRNAs were isolated by being labeled with BrUTP and immunoprecipitated with anti-BrU antibody, and then analyzed by microarray. More than 50 genes were identified as ZGA genes at 1-cell stage. The analysis of the regulatory regions of these genes revealed that a large part of them have the consensus sequences to which Pax and Ets family transcription factors bind.2. Analysis by a microarray of 1500 transcription factor genes showed that the expression patterns most changed between 1-cell and 2-cell stage and that the expression of Ets family transcription factors greatly increased after fertilization. Analysis for histone modifications by immunocytochemistny revealed that all modifications examined increased during growth of oocytes and that many modifications changed in various patterns during pre-plantation development.3. The growing oocytes were microinjected with RNAi and allowed to growth into full-grown oocytes, meiotic maturation and preimplantation development in vitro. We conformed that this system works well to knock-down a particular gene and analyze its function for the development of oocytes and preimplantaion embryos.