Genetic analysis of Kit ligand of mice

小鼠Kit配体遗传分析

• 批准号: 6729029
• 负责人: MARY A BEDELL
• 金额: $ 24.06万
• 依托单位: UNIVERSITY OF GEORGIA
• 依托单位国家: 美国
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-04-01 至 2007-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6729029
• 关键词: biological signal transduction; confocal scanning microscopy; cytokine; developmental genetics; gene interaction; gene mutation; glycosylation; immunoprecipitation; laboratory mouse; mammalian embryology; protein isoforms; protein structure function; protein tyrosine kinase; stem cell factor; tissue /cell culture; western blottings

DESCRIPTION (provided by applicant): The long-term goal of this research is to understand the mechanisms by which Kit ligand (Kitl) promotes the development of diverse cell types in mammnals. Kitl is the ligand for the Kit receptor tyrosine kinase and is a member of the short chain group of helical cytokines. In humans, Kit mutations cause gastrointestinal tumors, hematopoietic proliferative disorders and piebald trait. In mice, Kitl is encoded by the Steel (Sl) locus and studies of Kitl mutants have established that Kitl is essential to the embryonic and postnatal development of hematopoietic cells, germ cells and melanocytes. We propose to use a series of Kitl(sl) mutations, including eight newly characterized mutations, to genetically dissect Kitl function. Our previous studies have characterized the effects of these mutations on mouse survival, hematopoietic cells and germ cells and have identified mutations that exert strong and weak effects on the in vivo phenotype. We will use molecular and genetic approaches to determine how these mutations affect Kitl function and to gain a better understanding of the Kitl/Kit signaling pathway. In Specific Aim 1, we will use an in vitro system to examine the mechanism of altered function of these Kitl(sl) mutants. We will determine the intracellular processing and cell surface expression of Kitl(sl) mutants and determine if Kitl(sl) mutants are deficient for binding to Kit, for activation of Kit and for in vitro bioactivity: In Specific Aim 2, we will use comparative molecular modeling to predict the effects of Kitl mutations on Kitl structure. In Specific Aim 3, we will use selected Kitl(sl) mutants to dissect further the role of Kitl in three phases of germ cell development in embryos: during emergence of primordial germ cells (PGCs) from the epiblast; during migration of actively dividing PGCs of mid-gestation; and during the post-migratory phase of late gestation. In Specific Aim 4, we will genetically characterize a novel modifier of the Kitl about pigmentation phenotype in mice. By generating a new congenic strain, we will test whether this pigmentation modifier affects other aspects of the Kitl(sl) mutant phenotype. Ultimately, the gene responsible for this modifier will be identified and will provide new information about the Kitl signaling pathway. Together, these studies will provide new insights into Kitl function and may lead to novel strategies for the development of more effective cytokines.

描述(申请人提供)：本研究的长期目标是 了解Kit配体(Kitl)促进发育的机制 哺乳动物的不同细胞类型。Kitl是Kit受体的配体 酪氨酸激酶是螺旋细胞因子短链家族的一员。 在人类中，Kit突变会导致胃肠道肿瘤、造血 增殖性疾病和花斑特征。在小鼠中，Kitl由 Steel(Sl)基因座和Kitl突变体的研究已经证实Kitl是 对造血细胞的胚胎和出生后发育是必不可少的， 生殖细胞和黑素细胞。我们建议使用一系列Kitl(S1)突变， 包括8个新表征的突变，对Kitl进行基因解剖 功能。我们之前的研究已经表征了这些因素的影响 突变对小鼠存活、造血细胞和生殖细胞的影响 确定了对体内产生强影响和弱影响的突变 表型。我们将使用分子和遗传方法来确定这些 突变会影响Kitl的功能并更好地了解 Kitl/Kit信号通路。在具体目标1中，我们将使用体外系统 目的：研究这些Kitl(Sl)突变体功能改变的机制。我们会 确定Kitl(Sl)的细胞内加工和细胞表面表达 突变体，并确定Kitl(Sl)突变体是否缺乏与Kit结合的能力， 试剂盒的激活和体外生物活性：在特定目标2中，我们将使用 用比较分子模型预测Kit1基因突变对Kit1基因的影响 结构。在特定的目标3中，我们将使用选定的Kitl(Sl)突变体来解剖 此外，Kitl在胚胎生殖细胞发育的三个阶段中的作用： 在原始生殖细胞(PGC)从上胚层中出现的过程中；在 妊娠中期活跃分裂的原生殖细胞的迁移； 妊娠晚期的迁徙后阶段。在特定的目标4中，我们将从基因上 一种关于小鼠色素沉着表型的新的试剂盒修饰物的特征。 通过产生新的同源菌株，我们将测试这种色素沉着 修饰剂影响Kitl(Sl)突变表型的其他方面。最终， 负责这种修饰的基因将被鉴定出来，并将提供新的 有关Kitl信号通路的信息。总而言之，这些研究将 提供对Kitl功能的新见解，并可能导致新的战略 开发更有效的细胞因子。