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： 1．对1细胞阶段合子基因激活(ZGA)开始时表达的基因进行分析。2．通过转录因子和表观遗传修饰分析生长卵母细胞和早期植入前胚胎的基因表达调控。 3． RNAi 的功能分析这些研究的结果描述如下1。通过用 BrUTP 标记并用抗 BrU 抗体免疫沉淀来分离新生 mRNA，然后通过微阵列进行分析。超过50个基因在1细胞阶段被鉴定为ZGA基因。对这些基因调控区的分析表明，其中很大一部分基因具有Pax和Ets家族转录因子结合的共有序列。2．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细胞阶段变化最大，并且受精后Ets家族转录因子的表达大大增加。通过免疫细胞化学对组蛋白修饰的分析表明，所有检测到的修饰在卵母细胞生长过程中均有所增加，并且许多修饰在植入前发育过程中以各种模式发生变化。3．生长中的卵母细胞被显微注射 RNAi，并使其在体外生长为完全生长的卵母细胞、减数分裂成熟和植入前发育。我们确信该系统可以很好地敲低特定基因并分析其对卵母细胞和植入前胚胎发育的功能。