The Nodal Signaling Pathway In Embryonic Development

我们正在进行的节点信号分析涉及到条件诱变的使用：我们已经产生了一个条件“floxed”等位基因，它正在与表达Cre重组酶的转基因系在早期胚胎的各种谱系中一起使用。我们实验室对Nodal的其他工作是解决其表达的表观遗传调控。组蛋白修饰被认为是一种独立于其在染色质过程中的作用的表观遗传密码，组蛋白H3 （H3K4me3）上的三甲基化赖氨酸4主要存在于活性启动子区域，三甲基化赖氨酸27 （H3K27me3）标记被Polycomb蛋白抑制的基因。这两种修饰都存在于胚胎干细胞中重要发育基因的启动子中，最近被认为是一种表观遗传信号，标志着这些基因在发育后期的转录激活。我们已经发现了node - smad2 /3通路与Polycomb去抑制相关的证据。我们发现Nodal和Brachyury （Wnt/ β -连环蛋白信号在中胚层形成过程中的一个靶点，但我们现在已经确定它也是Nodal通路的一个直接靶点）都受到Polycomb抑制的调节。这两个基因都与Polycomb蛋白结合，在缺乏Nodal信号或Brachyury缺乏Nodal和Wnt信号的情况下显示H3K27me3抑制性组蛋白标记。我们进一步发现，node - smad2 /3信号通路直接参与H3K27me3去甲基化酶Jmjd3来抵消Polycomb在靶位点的抑制。Smads2/3与Jmjd3之间的物理相互作用以及Jmjd3向靶基因的募集依赖于活跃的信号传导。然而，在没有Polycomb功能的情况下，目标位点的表达独立于Nodal信号。这些结果确定Polycomb去抑制是node - smad2 /3通路的一种新功能，并表明这可能在干细胞和早期胚胎中起主要作用。

Our ongoing analysis of Nodal signaling involves the use of conditional mutagenesis: we have generated a conditional "floxed" allele, which is being used in conjunction with transgenic lines expressing Cre recombinase in various lineages of the early embryo. Additional work on Nodal in our lab is addressing the epigenetic regulation of its expression. Histone modifications have been proposed to function as an epigenetic code independent from their role in ongoing chromatin processes, with trimethylated lysine 4 on histone H3 (H3K4me3) mainly found in active promoter regions and trimethylated lysine 27 (H3K27me3) marking genes for repression by Polycomb proteins. The presence of both these modifications in promoters of developmentally important genes in embryonic stem (ES) cells recently has been suggested to function as an epigenetic signal marking these genes for transcriptional activation later in development. We have found evidence connecting the Nodal-Smad2/3 pathway with Polycomb de-repression. We have found that Nodal and Brachyury, a target of Wnt/beta-catenin signaling in mesoderm formation but which we have now identified as a direct target also of the Nodal pathway, are both regulated by Polycomb repression. Both genes are bound by Polycomb proteins and show H3K27me3 repressive histone marks in the absence of Nodal signaling, or in the absence of Nodal and Wnt signals in the case of Brachyury. We further found that the Nodal-Smad2/3 signaling pathway directly engages the H3K27me3 demethylase Jmjd3 to counteract Polycomb repression at target loci. Physical interaction between Smads2/3 with Jmjd3, and recruitment of Jmjd3 to target genes is dependent on active signaling. In the absence of Polycomb function, however, target loci are expressed independent of Nodal signaling. These results identify Polycomb de-repression as a novel function of the Nodal-Smad2/3 pathway, and suggest this may be a primary role in stem cells and perhaps the early embryo.