Isolation of Novel Mutations Affecting the Mouse Embryo

描述（由申请人提供）：组织哺乳动物胚胎身体计划的遗传途径和网络尚不清楚。在着床和妊娠中期之间，发生了轴决定、原肠胚形成、胚层特化、形态发生和器官系统的早期分化等关键事件，但调控这些过程的大多数基因尚未被鉴定。这是一个建议，继续使用乙基亚硝基脲（ENU）诱变，以确定基因，控制早期发育的小鼠胚胎。隐性突变之所以被识别是因为它们在妊娠中期胚胎中引起明显的形态异常。定位克隆用于鉴定负责发育表型的基因。这种方法已经确定了神经和中胚层模式所需的30个基因的突变，包括至少24个以前未表征的基因。已经确定了导致6种突变的分子病变，包括先前在小鼠中未研究的4种基因的突变。这项工作现在将集中在轴规范，图案和形态发生的早期事件。最近发现的影响早期模式和形态发生的突变将被绘制和分析。 影响这些过程的几个基因将被克隆和分子表征。为了提高控制早期植入后发育（包括轴特化和形态发生）的基因的鉴定效率，将产生在e7.5-e8.5胚胎中的限定细胞亚群中表达荧光蛋白的转基因系。这些报告细胞系将用于帮助表征已经鉴定的突变，并更有效地筛选影响早期模式和形态发生的新突变。新的突变将根据这些报告基因的不适当表达进行鉴定，并确定影响早期发育特定方面的基因组。 鉴定控制哺乳动物胚胎发育的基因对于检测、预防和治疗人类出生缺陷至关重要。在早期发育中起作用的信号通路在肿瘤发展中也起着关键作用，因此本研究中发现的这些基因可能会定义肿瘤发展和进展的新步骤。

DESCRIPTION (provided by applicant): The genetic pathways and networks that organize the body plan of the mammalian embryo are not well understood. Between implantation and midgestation, the critical events of axis determination, gastrulation, germ layer specification, morphogenesis and early differentiation of organ systems take place, but the majority of genes that regulate these processes have not yet been identified. This is a proposal to continue to use ethylnitrosourea (ENU) mutagenesis to identify genes that control early development in the mouse embryo. Recessive mutations are identified because they cause clear morphological abnormalities in the midgestation embryo. Positional cloning is used to identify the genes responsible for the developmental phenotypes. This approach has identified mutations in 30 genes required for neural and mesodermal patterning, including at least two dozen previously uncharacterized genes. The molecular lesions responsible for 6 mutations have been identified, including mutations in 4 genes that had not been studied previously in the mouse. The work will now focus on the early events of axis specification, patterning and morphogenesis. Recently identified mutations that affect early patterning and morphogenesis will be mapped and analyzed. Several genes that affect these processes will be cloned and characterized molecularly. To increase the efficiency of identification of genes that control early post-implantation development, including axis specification and morphogenesis, transgenic lines will be generated that express fluorescent proteins in defined subsets of cells in the e7.5-e8.5 embryo. These reporter lines will be used to help characterize mutations that have already been identified and to screen for new mutations that affect early patterning and morphogenesis more efficiently. New mutations will be identified based on inappropriate expression of these reporters and define groups of genes that affect specific aspects of early development. Identification of the genes that control development of the mammalian embryo is crucial for the detection, prevention and treatment of human birth defects. The signaling pathways that act in early development also have critical roles in tumor development, so it is likely that these genes identified in this study will define new steps in tumor development and progression.