TARGETING MUTATIONS OF TYROSINE KINASE GENES IN THE MOUSE

针对小鼠酪氨酸激酶基因的突变

• 批准号: 6989401
• 负责人: STEPHEN Paine GOFF
• 金额: $ 13.5万
• 依托单位: COLUMBIA UNIVERSITY HEALTH SCIENCES
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-04-12 至 2008-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6989401
• 关键词: B lymphocyte; alleles; biological signal transduction; cell cycle proteins; cell differentiation; cell line; developmental genetics; gene mutation; gene targeting; genetically modified animals; germ cells; laboratory mouse; mutant; oxidative stress; phenotype; protein localization; protein structure function; protein tyrosine kinase; protooncogene

This proposal describes experiments using the tools of mouse genetics to determine the in vivo functions of the c-abl proto-oncogene. The c-abl gene product is a cytoplasmic tyrosine kinase important for signal transduction and control of the cell cycle. Knock-out mice deficient in c-abl exhibit a number of phenotypes: perinatal lethality, runting, bone abnormalities, and defects in early lymphoid cell lineages. Several genetic studies of these mice are proposed. First, new alleles of c-abl will be generated by gene targeting ("knock-in" experiments). One of these alleles will permit the conditional deletion of the gene at selected times in development and in selected tissues; others will express altered forms of c-abl lacking particular domains responsible for protein localization. Examination of these mice should help determine the functions of each pathway emanating from c-abl; we will be particularly interested to examine the course of B cell development and the repertoire of variable region utilization. Secondly, a germ-line mutation of a new member of the Abi family termed PSTPIP1, a mammalian homologue of the yeast Cdc15 gene, will be generated by gene targeting, and the effects of the mutation alone and with other knock-out mutations will be characterized. Our expectation is that the loss of PSTPIP1 may suppress some of the phenotypes caused by loss of Abl. Thirdly, we will generate and analyze germ line mutations of the UV-DDB proteins, products of the xpe locus in man, and which we have shown are bound and regulated by the Abl kinase. Finally, we will directly catalogue the genes induced in cell lines and in tissues by oxidative stress, comparing the responses in abl mutants with wild-type controls, and thereby identify the Abl-dependent downstream targets. These studies should help define the many diverse functions of c-abl in mammalian development and physiology.

该提案描述了使用小鼠遗传学工具来确定 c-abl 原癌基因的体内功能的实验。 c-abl 基因产物是一种细胞质酪氨酸激酶，对于信号转导和细胞周期控制很重要。缺乏 c-abl 的基因敲除小鼠表现出多种表型：围产期致死、矮小、骨骼异常和早期淋巴细胞谱系缺陷。提出了对这些小鼠进行几项遗传学研究。首先，c-abl 的新等位基因将通过基因靶向（“敲入”实验）产生。这些等位基因之一将允许在发育中的选定时间和选定的组织中有条件地删除该基因；其他人会表达 c-abl 的改变形式，缺乏负责蛋白质定位的特定结构域。对这些小鼠的检查应有助于确定源自 c-abl 的每个通路的功能；我们将特别感兴趣的是检查 B 细胞的发育过程和可变区利用的全部功能。其次，Abi家族新成员PSTPIP1（酵母Cdc15基因的哺乳动物同源物）的种系突变将通过基因靶向产生，单独突变和与其他敲除突变的影响将是特点。 我们的预期是，PSTPIP1 的缺失可能会抑制 Abl 缺失引起的一些表型。第三，我们将生成并分析 UV-DDB 蛋白的种系突变，该蛋白是人类 xpe 基因座的产物，我们已证明该突变受 Abl 激酶的结合和调节。最后，我们将直接对细胞系和组织中氧化应激诱导的基因进行编目，比较 abl 突变体与野生型对照的反应，从而识别 Abl 依赖性下游靶标。这些研究应该有助于定义 c-abl 在哺乳动物发育和生理学中的多种不同功能。