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首页> 外文期刊>Molecular and Cellular Biology >Reconstitution of Enhancer Function in Paternal Pronuclei of One-Cell Mouse Embryos
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Reconstitution of Enhancer Function in Paternal Pronuclei of One-Cell Mouse Embryos

机译：重构单细胞小鼠胚胎父本核中的增强子功能

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How chromatin-mediated transcription regulates the beginning of mammalian development is currently unknown. Factors responsible for promoter repression and enhancer-mediated relief of this repression are not present in the paternal pronuclei of one-cell mouse embryos but are present in the zygotic nuclei of two-cell embryos. Here we show that coinjection of purified histones and a plasmid-encoded reporter gene into the paternal pronuclei of one-cell embryos at a specific histone-DNA concentration could recreate the behavior observed in two-cell embryos: acquisition of promoter repression and subsequent relief of this repression either by functional enhancers or by histone deacetylase inhibitors. Furthermore, the extent of enhancer-mediated stimulation in one-cell embryos depended on the acetylation status of the injected histones, on the treatment of embryos with a histone deacetylase inhibitor, and on the developmentally regulated appearance of enhancer-specific coactivator activity. The coinjected plasmids in one-cell embryos also exhibited chromatin assembly, as determined by a supercoiling assay. Thus, injection of histones into one-cell embryos faithfully reproduced the chromatin-mediated transcription observed in two-cell embryos. These results suggest that the need for enhancers to stimulate promoters through relief of chromatin-mediated repression occurs once the parental genomes are organized into chromatin. Furthermore, we present a model mammalian system in which the role of individual histones, and particular domains within the histones that are targeted in enhancer function, can be examined using purified mutant histones.
机译：染色质介导的转录如何调节哺乳动物发育的开始目前是未知的。负责启动子阻遏和增强子介导的这种阻抑的因素并不存在于单细胞小鼠胚胎的父本中，而存在于两细胞胚胎的合子核中。在这里，我们显示了在特定的组蛋白-DNA浓度下，将纯化的组蛋白和质粒编码的报告基因共注射到单细胞胚胎的父代前核中，可以重现在两细胞胚胎中观察到的行为：获得启动子抑制并随后缓解通过功能增强剂或组蛋白脱乙酰基酶抑制剂抑制。此外，单细胞胚胎中增强子介导的刺激程度取决于注射的组蛋白的乙酰化状态，组蛋白脱乙酰基酶抑制剂对胚胎的处理以及增强子特异性共激活子活性的发育调控外观。单细胞胚胎中的共注射质粒还显示出染色质组装，这是通过超螺旋测定法确定的。因此，将组蛋白注射到单细胞胚胎中可以忠实地复制在两细胞胚胎中观察到的染色质介导的转录。这些结果表明，一旦亲本基因组被组织成染色质，就需要增强剂通过减轻染色质介导的阻抑来刺激启动子。此外，我们提出了一种模型哺乳动物系统，其中可以使用纯化的突变组蛋白检查单个组蛋白的作用，以及组蛋白内针对增强子功能的特定结构域的作用。

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