Detection of BMP Signaling In Pre-Implantation Mouse Embryos

描述(申请人提供)：哺乳动物的发育始于受精，随后是多个细胞分裂事件，以及从母体转录到合子转录的转变。所有这些事件协同作用，建立了第一个也是非常重要的发育事件--血统分化。在植入前胚胎建立不同谱系的第一步是滋养外胚层(TE)和内细胞团(ICM)，滋养外胚层是胎盘和卵黄囊等胚外结构的前体，内细胞团成为胚胎本身。虽然已经广泛研究了Oct3/4、Nanog、Sox2、Tead4和CDx2等转录因子的调节相互作用和形态贡献，但令人惊讶的是，对分泌生长因子在体内的作用知之甚少。 这些发展的早期阶段。骨形态发生蛋白(BMP)在这些早期小鼠胚胎阶段的存在表明BMP信号在谱系分化中可能发挥功能作用。本项目将通过阐明BMP信号的时空出现和功能作用来确定BMP信号在植入前胚胎中的意义。此外，这个项目的目的是确定BMPs和谱系规范转录因子之间的分子相互作用，这些转录因子对第一次谱系分化至关重要。通过一种高灵敏度的图像分割定量方法，免疫荧光检测和定量磷酸化的Smad1/5/8将表明植入前胚胎中活跃的BMP信号。功能丧失实验将被用来确定BMP信号活性是否影响到在胚泡形成过程中控制谱系指定的两个主要转录网络：Oct4/Nanog/Sox2的多功能调控基序和Tead4、CDX2和GATA6的TE提交网络。BMP信号在着床前哺乳动物胚胎发育中的生物学作用的直接证据为理解转录调控因子和信号形态原之间的新的调控相互作用提供了基础。这一理解有利于多个与健康相关的学科，如干细胞治疗学、癌症生物学和生殖医学。

DESCRIPTION (provided by applicant): Mammalian development begins with fertilization, followed by multiple cell cleavage events and the transition from maternal to zygotic transcription. All these events work in concert to establish the first and thus highly significant developmental event, lineage differentiation. The first step in the establishment of different lineages in the pre-implantation embryo gives rise to the trophectoderm (TE), the precursor for extraembryonic structures such as the placenta and yolk sac, and the inner cell mass (ICM), which becomes the embryo proper. While the regulatory interactions and morphological contributions of transcription factors such as Oct3/4, Nanog, Sox2, Tead4, and Cdx2 have been extensively studied, surprisingly little is known about the roles of secreted growth factors during these early stages of development. The presence of bone morphogenetic proteins (BMPs) in these early mouse embryonic stages suggests likely functional roles for BMP signaling in lineage differentiation. This project will determine the significance of BMP signaling in pre- implantation stage embryos by elucidating the spatiotemporal emergence and functional role of BMP signaling. Also, this projects aims to identify molecular interactions between BMPs and lineage specification transcription factors vital for the first lineage differentiation. Through a highly sensitive image segmentation quantification method, immunofluorescence detection and quantification of phosphorylated Smad1/5/8 will denote active BMP signaling in pre-implantation embryos. Loss of function experiments will be used to determine if BMP signaling activity impinges on two major transcriptional networks that control lineage specification during blastulation: the pluripotent regulatory motifs of Oct4/Nanog/Sox2 and the TE committing networks of Tead4, Cdx2, and Gata6. Direct evidence for the biological role of BMP signaling in pre-implantation mammalian embryonic development provides a foundation for understanding novel regulatory interactions between transcriptional regulators and signaling morphogens. This understanding benefits multiple health related disciplines such as stem cell therapeutics, cancer biology, and reproductive medicine.