FGD1/CDC42: SIGNAL TRANSDUCTION & DEVELOPMENTAL GENETICS

FGD1/CDC42：信号转导

• 批准号: 2674010
• 负责人: JEROME L. GORSKI
• 金额: $ 30.87万
• 依托单位: UNIVERSITY OF MICHIGAN AT ANN ARBOR
• 依托单位国家: 美国
• 财政年份: 1997
• 资助国家: 美国
• 起止时间: 1997-09-09 至 2001-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2674010
• 关键词: biological signal transduction; developmental genetics; embryo /fetus cell /tissue; gene expression; gene induction /repression; gene mutation; gene targeting; genetic disorder; genetically modified animals; guanine nucleotide binding protein; guanine nucleotide exchange factors; guanosinetriphosphatases; histogenesis; immunocytochemistry; laboratory mouse; microinjections; model design /development; molecular cloning; polymerase chain reaction; protein kinase; sex linked trait; skeletal disorder; stress proteins; vertebrate embryology; yeast two hybrid system; zebrafish

DESCRIPTION: This research aims to analyze the molecular genetics of faciogenital dysplasia (FGDY), an X-linked developmental disorder that adversely affects the formation of multiple skeletal structures including elements of the face, spine, and distal extremities. The gene responsible for this disorder, FGD1, encodes a guanine nucleotide exchange factor (GEF) that specifically activates Cdc42, a member of the Rho (Ras-homology) family of the p21 GTPases. By activating Cdc42, FGD1 stimulates fibroblasts to form filopodia, cytoskeletal elements involved in cellular signaling, adhesion and migration. Through Cdc42, FGD1 also activates the stress-activated protein kinase (SAPK) signaling cascade, a pathway that regulates cell growth and differentiation. Preliminary studies show that FGD1 is primarily expressed in skeletal progenitors including vertebral precartilagenous condensations, ossifying cervical vertebrae, and ossifying craniofacial bones, bones affected in FGDY. However, data also shows that mammalian genomes contain multiple FGD1 homologues, a result that suggests that FGD1 may be a component of a partially redundant Cdc42 activation system. Together, the accumulated data indicates that FGD1 is a component of a developmentally regulated signaling pathway involved in cellular morphology and mammalian skeletogenesis. The overall goals of this application are to develop a comprehensive model to further elucidate the role of the FGD/Cdc42 signaling cascade in mammalian morphogenesis. To this end, cross-hybridization and degenerate PCR amplification techniques will be used to isolate additional FGD1 homologues; mouse and zebrafish FGD cDNA clones will be used to determine the spatial and temporal patterns of gene expression during embryogenesis. To provide precise cellular localization, FGD1-directed antibodies will be used to study embryonic tissue by immunocytochemistry. Microinjection and in vitro assays will be used to study the molecular biology and to molecularly dissect the functional domains of the FGD proteins. Two-hybrid and other molecular techniques will be used to isolate and characterize additional components of the FGD1/Cdc42 signal transduction pathway. Transgenic mice that express recombinant FGD1 proteins will be used to study the developmental consequences of dysregulated FGD1 expression. FGD-deficient mice will be generated to study the developmental consequences of null FGD genotypes. Compound FGD-deficient mice will be generated to examine FGD homologues for functional redundancy. These analyses will provide a means to study the molecular genetics of the FGD/Cdc42 signal transduction pathway in mammalian morphogenesis. These analyses should result in a more thorough understanding of skeletogenesis, signal transduction, and the general principles responsible for the diversity of skeletal form.

描述：本研究旨在分析黑色素瘤的分子遗传学。 面性发育不良(FGDY)，一种X连锁的发育障碍， 对多种骨骼结构的形成产生不利影响，包括 面部、脊柱和四肢远端的元素。负责的基因 对于这种疾病，FGD1编码一种鸟核苷酸交换因子(Egf)。 它特异性地激活了Rho(RAS同源)家族的成员CDC42 P21GTP酶。通过激活CDC42，FGD1刺激成纤维细胞 形成丝状伪足，参与细胞信号传递的细胞骨架元素， 黏附和迁移。通过CDC42，FGD1还激活了 应激激活蛋白激酶(SAPK)信号转导通路 调节细胞的生长和分化。初步研究表明， FGD1主要在骨骼祖细胞中表达，包括脊椎 软骨前浓缩、颈椎骨化和骨化 颅面部骨骼，受FGDY影响的骨骼。不过，数据也显示， 哺乳动物基因组包含多个FGD1同源物，这一结果表明 FGD1可能是部分冗余的CDC42激活的组成部分 系统。总而言之，累积的数据表明FGD1是一个组件 一种发育调节的信号通路参与细胞 形态和哺乳动物的骨骼发生。 该应用程序的总体目标是开发一个全面的模型 为了进一步阐明FGD/CDC42信号级联在细胞内的作用 哺乳动物的形态发生。为此，杂交和退化 将使用聚合酶链式反应扩增技术分离出更多的FGD1 同源基因；小鼠和斑马鱼FGD基因克隆将用于确定 胚胎发育过程中基因表达的时空模式。 为了提供精确的细胞定位，FGD1定向的抗体将 用于通过免疫细胞化学研究胚胎组织。微量注射和 体外试验将被用于研究分子生物学和 从分子水平上剖析FGD蛋白的功能结构域。双杂交 和其他分子技术将被用来分离和鉴定 FGD1/CDC42信号转导通路的其他组件。 表达重组FGD1蛋白的转基因小鼠将用于研究 FGD1表达异常的发育后果。 将产生FGD缺陷小鼠来研究其发育后果 FGD缺失型。复合FGD缺陷小鼠将产生 检查FGD同系物的功能冗余。这些分析将 为研究FGD/Cdc42信号的分子遗传学提供了一种手段 哺乳动物形态发生中的转导途径。这些分析应该 导致对骨骼形成、信号 转导，以及导致基因多样性的一般原则。 骨瘦如柴。