Phenotyping Early Embryonic Lethal Knockout Mice to Identify Essential Genes with Previously Uncharacterized Roles in Pre-implantation Development, Gastrulation, Turning, and Placentation

对早期胚胎致死性基因敲除小鼠进行表型分析，以确定在植入前发育、原肠胚形成、转向和胎盘着床中具有先前未表征的作用的重要基因

基本信息

批准号：
9906060
负责人：
ANNA-KATERINA HADJANTONAKIS
金额：
$ 68.91万
依托单位：
SLOAN-KETTERING INST CAN RESEARCH
依托单位国家：
美国
项目类别：
财政年份：
2016
资助国家：
美国
起止时间：
2016-06-10 至 2021-04-30
项目状态：
已结题

项目摘要

DESCRIPTION (provided by applicant): Advances in genomic technologies over the past decade have yielded an unprecedented level of information about animal genomes. However, much of the information underlying these 2D models of cell and tissue function stills awaits experimental verification and further investigation in intact living organisms. Another factor limiting the application of genome wide approaches is the absence of functional characterization for ≥ 1/2 of all annotated genes. The targeted mutations generated by the International Knockout Mouse Consortium (IKMC), including KOMP, provide a revolutionary resource for functionally annotating the mammalian genome. We propose to characterize the phenotypes of mutations in the KOMP2-generated KO lines that result in develop- mental arrest or abnormal morphology at or prior to E9.5. We will functional define ~200 previously uncharacterized genes whose functions are essential during pre- and early post-implantation development. The investigators of this group have previously studied the phenotypes of ~200 embryonic lethal mouse mutants, leading to many surprising findings, including the novel cellular mechanisms that create the mammalian endoderm, the requirement for primary cilia in the Hedgehog signaling pathway and many more. Our specific goals are to (1) Characterize the functions of new genes essential for early embryonic development, by second tier phenotyping of ≥200 lines lethal at or before e9.5. We will define the stage of arrest, analyze morphological abnormalities, assess proliferation and cell death. Only genes in which knock- outs have not been characterized will be studied, and those encoding uncharacterized proteins will have highest priority, making this an unbiased screen for novel essential genes, and providing the first evidence on bio- logical function of 200 essential genes. (2) Use our expertise to characterize at cellular resolution the roles of previously uncharacterized genes in preimplantation development, early embryonic morphogenesis and placental development. We will characterize novel regulators of pluripotency of early embryonic lineages, the gastrulation epithelial-mesenchymal transition, early mesoderm migration and ventral folding, which are reiteratively used morphogenetic programs essential for many aspects of human development. Characterization of cellular and developmental functions of regulators of placentation is crucial for fetal and child health. Our specific Aims are: Aim 1. - Establishing a Phenotyping Pipeline, which will initiate at three trans NIH KOMP2 production and phenotyping centers. Aim 2 - Tier two phenotyping of ~200 mutations that cause lethality before E9.5. Aim 3 - Tier 3 phenotyping of lethal mutations that affect development of the blastocyst, early post-implantation morphogenesis and placentation. The embryonic lethal KO mutations generated by KOMP and IKMC provide an unprecedented opportunity to expand and enrich the functional annotation of the mammalian genome. The embryonic lethal genes thus identified will not only be indispensable for early mouse development, but also critical for multiple aspects of human development and tissue homeostasis.

描述（通过应用程序提供）：过去十年中基因组技术的进步已经产生了有关动物基因组的前所未有的信息。但是，这些2D模型的细胞和组织功能模型静止的信息静止不动，这些信息正在等待实验验证，并在完整的生物体中进行进一步研究。限制基因组广泛方法应用的另一个因素是所有注释基因的1/2≥1/2的功能表征。包括KOMP在内的国际基因敲除小鼠财团（IKMC）产生的有针对性突变为在功能上注释哺乳动物基因组提供了革命性的资源。我们建议表征KOMP2生成的KO系中突变的表型，这些突变在E9.5或之前或之前导致心理停滞或异常形态。我们将功能定义约200个以前未表征的基因，其功能在植入前和早期植入后发育中至关重要。该组的研究人员先前已经研究了约200个胚胎致死小鼠突变体的表型，从而导致许多令人惊讶的发现，包括创建哺乳动物内胚层的新型细胞机制，在刺猬信号通路中对原代纤毛的需求等等。我们的具体目标是（1）通过在E9.5或之前或之前通过第二层表型致死的第二层表型来表征早期胚胎发育必不可少的新基因的功能。我们将定义逮捕阶段，分析形态异常，评估增殖和细胞死亡。只有尚未表征敲除的基因才能进行研究，而编码未表征蛋白质的人将具有最高优先级，这使它成为新基本基因的无偏筛选，并提供了有关200个必需基因的生物逻辑功能的第一个证据。（2）利用我们的专业知识来表征细胞分辨率的表征，以前未表征基因在植入前发育，早期胚胎形态发生和占地发育中的作用。我们将表征早期胚胎谱系多功能的新型调节剂，胃上皮 - 间质转变，早期中胚层迁移和腹侧折叠，这是对人类发展许多方面必不可少的形态发育程序。调节剂的细胞和发育功能的表征对于胎儿和儿童健康至关重要。我们的具体目的是：目标1。-建立一个适用于型管道，该管道将在三个跨性别 NIH KOMP2生产和表型中心。目标2- 〜200个突变的两个表型在E9.5之前引起致死性。 AIM 3 -TIER 3的致命突变表型，影响胚泡的发展，早期植入后形态发生和位置。 KOMP和IKMC产生的胚胎致死性KO突变为扩展和丰富哺乳动物基因组的功能注释提供了前所未有的机会。因此，鉴定出的胚胎致死基因不仅对于早期小鼠发育是必不可少的，而且对于人类发育和组织稳态的多个方面也至关重要。