Genome-wide ENU mutagenesis screen for Runx2 modifiers

Runx2 修饰符的全基因组 ENU 诱变筛选

• 批准号: 6956298
• 负责人: Patricia Florence Ducy
• 金额: $ 9.75万
• 依托单位: BAYLOR COLLEGE OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-08-01 至 2006-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6956298
• 关键词: X ray; bioassay; biotechnology; bone development; bone development disorder; cell differentiation; developmental genetics; gene mutation; genetic mapping; genetic screening; laboratory mouse; mesenchyme; molecular cloning; nitrosourea; osteoblasts; phenotype; structural genes; transcription factor

DESCRIPTION (provided by applicant): The long-term goal of our project is to identify factors regulating mesenchymal cells commitment and early maturation into differentiated osteoblasts. As a first step toward this goal, we propose to perform a genome-wide genetic screen in mice using ethyl-nitrosourea (ENU) mutagenesis to expose mutations able to functionally compensate for Runx2 haploinsufficiency. Runx2 is the earliest known determinant of osteoblast differentiation, and loss of one Runx2 allele in mice or human causes Cleidocranial dysplasia (CCD), a skeletal disorder marked by clavicle agenesis and ossification defects. Any regulator of Runx2 expression or activity is thus likely to be involved in controlling either osteoblast differentiation or osteoblast function. Our strategy relies on a search for ENU-mutated animals that harbor a rescued CCD phenotype despite a Runx2 genetic background. We hypothesize that such rescuing mutations will affect regulators of Runx2 gene expression or protein stability/availability/activity or trigger yet unknown parallel or downstream compensating pathways. Subsequent characterization of the genes harboring these mutations should thus identify regulators involved in the early steps of skeletogenesis. The present application proposes to identify rescued Runx2 ENU mutants and to select the best candidates for subsequent mutation mapping and gene identification. Our specific aims are: to identify ENU-mutated mouse strains harboring a rescue of the CCD phenotype despite heterozygocity at the Runx2 locus; and to compare the mutant lines isolated and prioritize them for subsequent mutation mapping.

描述（由申请人提供）：我们项目的长期目标是确定调节间充质细胞承诺和早期成熟为分化成骨细胞的因素。作为朝着这一目标的第一步，我们建议使用乙基硝化乙烯（ENU）诱变在小鼠中进行全基因组遗传筛选，以暴露能够在功能上补偿Runx2单倍弥补的突变。 Runx2是成骨细胞分化的最早的决定因素，并且在小鼠或人类引起的一个Runx2等位基因的损失中丧失了裂解症状增生症（CCD），这是一种骨骼疾病，标志着锁骨发育不全和骨化缺陷。因此，Runx2表达或活性的任何调节剂都可能参与控制成骨细胞分化或成骨细胞功能。我们的策略依靠寻找具有Runx2遗传背景的ENU突变动物，该动物具有救出的CCD表型。我们假设这种救援突变会影响Runx2基因表达或蛋白质稳定性/可用性/活动或触发且未知的平行或下游补偿途径的调节剂。因此，随后的具有这些突变的基因的表征应确定涉及骨骼生成早期步骤的调节剂。本应用建议识别救出的Runx2 ENU突变体，并选择最佳的候选物，以进行随后的突变映射和基因鉴定。我们的具体目的是：确定尽管在Runx2基因座的杂合性，但仍具有CCD表型的ENU突变的小鼠菌株；并比较分离的突变线并确定其优先级以进行随后的突变映射。