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

卵母细胞和植入前胚胎基因表达重编程机制

基本信息

批准号：
17380166
负责人：
AOKI Fugaku
金额：
$ 9.04万
依托单位：
The University of Tokyo
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
2005
资助国家：
日本
起止时间：
2005 至 2007
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-17380166/
关键词：
oocyte embyro gene exuression reprogramming histone ヒストン修飾発生受精減数分裂

项目摘要

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.

本研究项目包括以下3个步骤.在1-细胞阶段合子基因激活（ZGA）开始时表达的基因的概况。通过检测转录因子和表观遗传修饰分析生长期卵母细胞和早期植入前胚胎的基因表达调控.这些研究的结果描述如下1.用BrUTP标记新生的mRNA，用抗BrU抗体免疫沉淀，然后用微阵列分析。在1-细胞期鉴定出50多个ZGA基因。对这些基因调控区的分析表明，它们中的很大一部分具有与Pax和Ets家族转录因子结合的共有序列.通过对1500个转录因子基因的微阵列分析表明，表达模式在1-细胞和2-细胞阶段之间变化最大，并且Ets家族转录因子的表达在受精后大大增加。免疫细胞化学分析组蛋白修饰显示，在卵母细胞的生长过程中，所检测的组蛋白修饰均增加，并且在种植前发育过程中，许多组蛋白修饰以不同的模式发生变化.将生长中的卵母细胞用RNAi显微注射，并允许其在体外生长成完全生长的卵母细胞、减数分裂成熟和植入前发育。我们证实，该系统可以很好地敲除特定的基因，并分析其对卵母细胞和预受精胚胎发育的功能。