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Genetic analysis of Kit ligand of mice

小鼠Kit配体遗传分析

基本信息

批准号：
7036715
负责人：
MARY A BEDELL
金额：
$ 23.75万
依托单位：
UNIVERSITY OF GEORGIA
依托单位国家：
美国
项目类别：
财政年份：
2002
资助国家：
美国
起止时间：
2002-04-01 至 2008-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7036715
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

项目摘要

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（由申请人提供）：本研究的长期目标是了解Kit配体（Kitl）促进发展的机制不同类型的细胞。Kit 1是Kit受体的配体酪氨酸激酶，并且是螺旋细胞因子短链组的成员。在人类中，Kit突变会导致胃肠道肿瘤、造血系统肿瘤、增殖性疾病和花斑性状。在小鼠中，Kitl由 Steel（Sl）基因座和Kitl突变体的研究已经确定Kitl是对造血细胞的胚胎和出生后发育至关重要，生殖细胞和黑素细胞。我们建议使用一系列Kitl（sl）突变，包括八个新发现的突变，功能我们以前的研究已经描述了这些因素的影响，突变对小鼠存活、造血细胞和生殖细胞的影响，鉴定出的突变对体内的表型我们将使用分子和遗传学方法来确定这些突变影响Kitl功能，为了更好地了解 Kitl/Kit信号通路。在具体目标1中，我们将使用体外系统以检查这些Kitl（sl）突变体的功能改变的机制。我们将确定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的结合，试剂盒活化和体外生物活性：在特定目标2中，我们将使用比较分子建模以预测Kitl突变对Kitl的影响结构在特定目标3中，我们将使用选择的Kitl（sl）突变体来解剖进一步研究了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（PGCs）从外胚层出现期间;在妊娠中期活跃分裂的PGCs的迁移; 后迁移阶段的晚期妊娠。在第四个目标中，我们将从基因上描述了一种关于小鼠色素沉着表型的Kitl的新修饰剂。通过产生新的同类菌株，我们将测试这种色素沉着是否修饰子影响Kitl（sl）突变体表型的其它方面。最后，负责这种修饰的基因将被鉴定出来，关于Kitl信号通路的信息。这些研究将为Kitl功能提供新的见解，并可能导致新的策略，开发更有效的细胞因子。