FUNCTION OF FIBROBLAST GROWTH FACTOR GENE IN MAMMALIAN EMBRYOGENESIS

成纤维细胞生长因子基因在哺乳动物胚胎发生中的功能

• 批准号: 5213815
• 负责人: GAIL R MARTIN
• 金额: --
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/5213815
• 关键词: biological signal transduction; chick embryo; developmental genetics; early embryonic stage; embryonic stem cell; fibroblast growth factor; gene expression; gene targeting; genes; genetic manipulation; genetic recombination; genetically modified animals; growth factor receptors; histogenesis; immunocytochemistry; in situ hybridization; laboratory mouse; mammalian embryology; mesoderm; molecular cloning; monoclonal antibody; nucleic acid probes; polymerase chain reaction; transfection

Evidence is rapidly accumulating that basic fibroblast growth factor 9b- FGF) and related proteins play an important role in embryonic induction. In mammals, four genes (a-FGF, k-FGF, int-2 and FGF-5) have been identified that have extensive sequence similarity to b-FGF, and more are likely to exist. We are interested in studying the role of these genes in early mammalian development. Our first set of specific goals is to determine when and where each of these genes is expressed in the developing mouse embryo by in situ RNA hybridization and protein localization, and from this information to begin to build a hypothesis about what their function in development might be. However, since the stage- and tissue-specific functions of these different growth factors will depend on the stage- and tissue- specificity of their respective receptors, we also propose to study the expression of FGF-receptor(s) in the developing embryo. In order to achieve these goals, we must have available probes for each of these mouse genes. Thus far, the only mouse gene in the FGF family that has been cloned in int-2. We have employed a variety of techniques that utilize the polymerase chain reaction (PCR) to clone cDNAs for all the known members of the mouse FGF family and have begun to preliminary examination of their expression during embryonic development. We are extending these studies to isolate hitherto unknown members of the family. In collaboration with Dr. L. Williams group, we propose to use similar approaches to obtain the mouse FGF-receptor (FGF-R) genes. As these different genes become available, we will carry out a detailed study of their expression in the developing embryo. As information on the expression of these genes begins to accumulat, it should be possible to infer something about their respective functions in development, some may display patterns of expression in the developing embyo. As information on the expression of these genes begins to accumulate, it should be possible to infer something about their respective functions in development, some may display patterns of expression consistent with a role in development of the embryonic heart or lungs. We should be aided in drawing these inferences by frequent discussions with Dr. M. Kirschner and his collaborators, who are studying the role of b-FGF in mesoderm induction in the frog. Our second set of specific goals is to obtain mice carrying mutations in the specific FGF or FGF-R genes of interest. This will be achieved by transfecting mouse embryonic stem (ES) cells in vitro with altered copies of the gene of interest and selecting those cell in which the altered allele has replaced an endogenous copy of the gene by homologous recombination. Such cells would then be used to generate mouse strains heterozygous for the altered genes. These mice could then be inter-bred to produce offspring homozygous for the targeted genetic change, and the functional consequences of the genetic alterations assessed. The results of such studies should allow us to evaluate any hypotheses we have developed on the possible funcion of the FGF and FGF-R gene familes in development.

证据正在迅速积累，碱性成纤维细胞生长因子9 b- FGF）及其相关蛋白在胚胎诱导中起重要作用。 在哺乳动物中，四种基因（a-FGF、k-FGF、int-2和FGF-5）已被发现。 鉴定出与b-FGF具有广泛的序列相似性， 可能存在。我们有兴趣研究这些基因在 哺乳动物早期发育 我们的第一组具体目标是确定何时何地 这些基因通过原位RNA在发育中的小鼠胚胎中表达， 杂交和蛋白质定位，并从这些信息， 开始建立一个假设， 来说也许是不对的然而，由于阶段和组织特异性的功能， 这些不同的生长因子将取决于阶段和组织， 它们各自受体的特异性，我们还建议研究 FGF受体在发育胚胎中的表达。为了 为了实现这些目标，我们必须为每一个目标提供可用的探测器。 老鼠基因到目前为止，FGF家族中唯一的小鼠基因 在int-2中被克隆。我们采用了多种技术， 利用聚合酶链反应（PCR）克隆所有的cDNA， 小鼠FGF家族的已知成员，并已开始初步研究 研究它们在胚胎发育过程中的表达。我们 将这些研究扩展到分离迄今未知的 家人与L博士合作。威廉姆斯集团，我们建议使用 类似的方法来获得小鼠FGF-受体（FGF-R）基因。作为 这些不同的基因变得可用，我们将进行详细的 研究它们在发育中的胚胎中的表达。作为关于 当这些基因的表达开始积累时， 推断出它们各自在发育中的作用，有些人可能 在发育中的胚胎中显示出表达模式。的资料 当这些基因的表达开始积累时， 为了推断它们各自在发育中的作用，一些 可能显示出与发育中的作用一致的表达模式 心脏或肺的胚胎。我们应该得到帮助， 通过与M博士的频繁讨论进行推断。Kirschner和他的 研究b-FGF在中胚层诱导中的作用的合作者 在青蛙。 我们的第二组具体目标是获得携带突变的小鼠， 目的特定FGF或FGF-R基因。为实现这些目标将 小鼠胚胎干细胞的体外复制 并选择其中改变的基因的那些细胞， 等位基因已被同源基因取代了基因的内源性拷贝， 重组这些细胞将被用来产生小鼠品系 变异基因的杂合子这些老鼠可以进行杂交 以产生对于靶向遗传变化纯合的后代，并且 评估遗传改变的功能后果。结果 这样的研究应该让我们评估任何假设，我们有 研究FGF和FGF-R基因家族在人乳腺癌中的可能功能， 发展