Tecto调节非洲爪蛙胚层决定与分化的机制研究

The cell fate determination during embryonic development of multicellular organisms involves both extrinsic and intrinsic regulators. The cells in the animal pole of Xenopus blastula will be eventually specified into ectoderm. However these cells can be converted into mesoderm or endoderm during balstula satges and are not fully specified into ectoderm unitl mid-gastrula stage. Whether the ectodermal fate is regulated by maternal factors is largely unknown. Recently, it was reported that the somatic type linker histone H1 and the noncanonical variant of histone H3 (H3.3) are involved in the competence of the cells in the animal pole to mesoderm induction. The optimal ratio of H1 and H3.3 is critical for the mesoderm induction competence. In our preliminary experiments, we found that a novel maternal transcriptional factor Tecto, a homolgue of VegT， is necessary and sufficient for ectoderm specification. We also found ectopic expression of Tecto inhibited the formation of mesoderm in during gastrula stages, and the competence of animal cells to exogenous and endogenous mesoderm inducing signals. The proposed research will determine whether Tecto is able to regulate the depostion of H1 and H3.3 in gene loci that are critical for germ layer formation. We hypothesize that Tecto may regulate the relative abundance of H1 and/or H3.3 in the regulatory regions of germ layer formation and differentiation genes.

动物胚胎发育期间细胞命运的决定受环境中的诱导信号和细胞本身的状态或者响应性有关。非洲爪蛙囊胚期动物极细胞的预定命运是外胚层的衍生物：表皮和神经。但是在原肠胚中期以前动物极细胞可以响应多种生长因子的诱导而转分化成中胚层或者内胚层；之后这些动物极细胞获得稳定外胚层发育命运。囊胚期中、内胚层的生成均受母源因子VegT的调控，但是外胚层是否由母源因子的控制还不得而知。文献报道合子型组蛋白H1与H3.3在染色质组装过程中的掺入比例与与囊胚期动物极细胞对中、内层诱导的响应性有关，但是其调控机制还不清楚。我们的前期实验发现与VegT同源的母源转录因子Tecto具有促进外胚层、抑制中、内胚层发育的功能。敲降母源Tecto的储备，导致外胚层基因表达下调和胚胎发育缺陷。我们将通过系列遗传、生化以及组学方法检验在囊胚期动物极细胞中Tecto是否能够调节H1和H3.3在胚层发育相关基因位点上的相对丰度。

胚层决定和分化是动物胚胎早期发育研究的关键问题之一。两栖类模式动物非洲爪蛙囊胚期外胚层同哺乳类囊胚期上胚层类似，具有多潜能。本课题解析一个新的关键母源因子Ascl1在调控非洲爪蛙囊胚期胚层决定和分化中的作用及机理。取得以下两项主要研究成果：.1、经典的促神经因子Ascl1作为母源因子在胚层决定和分化中具有双重活性，作为转录激活因子赋予囊胚期外胚层的神经分化倾向，作为转录抑制因子拮抗母源中内胚层决定因子VegT的活性，从而决定囊胚期胚层命运的分配。发表两篇相关研究论文。.2、Ascl1的氨基端在进化上高度保守，能够招募HDAC1，并与VegT相互作用，调控中内胚层基因启动子区域H3K27ac/H3K9ac表达水平，进而决定中内胚层基因的表达强度。.在以上两项研究成果的基础上，我们还探索了母源Ascl1调控外胚层的神经分化潜能的分子机制，获得了多个Ascl1的靶基因，为进一步深入研究囊胚期外胚层的决定和分化机制奠定基础。