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Gene expression in early mouse embryo and its regulation

小鼠早期胚胎基因表达及其调控

基本信息

批准号：
11694224
负责人：
KIMURA Minoru
金额：
$ 6.46万
依托单位：
Tokai University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
1999
资助国家：
日本
起止时间：
1999 至 2001
项目状态：
已结题

项目摘要

In early embryogenesis of various species, there is a transition of matemal RNA to zygotic RNA, which is thought to play an important role in molecular programming of later stage of development. We first found in mice that one type of maternal RNA, which was first isolated in our laboratory and termed "SSEC-D(stage-specific embryonic clone-D)", exhibited a transient extension of poly (A) strand at its 3' end with a peak at 18h after fertilization (corresponding to the late 1-cell stage) and a gradual degradation of the poly(A) strand thereafter. Based on this finding, we have examined this phenomenon in more detail in the past three years, and obtained several findings as mentioned below.1. Transient increase in the size of SSEC-D RNA is due to addition of poly (A) strand at its 3' end, and it requires new protein synthesis. 2. It is possible to perform Northern blot analysis using 10-20 zygotes and early embryos due to improvement of mRNA detection system. 3. Transient extension of RNA was also observed in zygotes after cytoplasmic injection experiments using in vitro synthesized RNAs containing EGFP cDNA ligated to various sizes of 3'-noncoding region of SSEC-D RNA. Thus, this phenomenon is already programmmed. 4. Addition of poly(A) spanning 300-400 nucleotides can release the translationally suppressed state of the maternal RNA iteself. 5. This poly(A) extension is dependent on the sequence of SSEC-D RNA itself and poly(A) sequence at its 3' end region. 6. It becomes possible to concentrate cDNAs enriched with maternal RNAs by improving the uptake of biotin labeled ATP by zygotes.

在不同物种的早期胚胎发生中，存在着从母体RNA到合子RNA的过渡，这被认为在发育后期的分子编程中起着重要的作用。我们首次在小鼠中发现了一种母体RNA，命名为SSEC-D(阶段特异性胚胎克隆-D)，它在受精后18h(对应于1-细胞晚期)显示出多(A)链的瞬时延伸，并在此后逐渐降解。基于这一发现，我们在过去三年中对这一现象进行了更详细的研究，并获得了如下所述的几个发现。SSEC-D RNA大小的暂时性增加是由于其3‘端添加了聚(A)链，需要新的蛋白质合成。2.由于mRNA检测体系的改进，利用10-20个受精卵和早期胚胎进行Northern杂交分析成为可能。3.体外合成的含有不同大小的SSEC-D RNA 3‘-非编码区的EGFP基因的RNA经细胞质注射实验后，受精卵中也观察到了RNA的瞬时延伸。因此，这种现象已经被程序化了。4.添加跨越300-400个核苷酸的聚(A)可以释放母体RNA本身的翻译抑制状态。5.这种多聚(A)延伸依赖于SSEC-D RNA本身的序列及其3‘末端的多聚(A)序列。6.通过提高受精卵对生物素标记的ATP的摄取，可以浓缩富含母体RNA的cDNA。