FGD1/CDC42: SIGNAL TRANSDUCTION & DEVELOPMENTAL GENETICS

FGD1/CDC42：信号转导

• 批准号: 2409016
• 负责人: JEROME L. GORSKI
• 金额: $ 28.68万
• 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/2409016
• 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 面生殖器发育不良（FGDY），一种X连锁发育障碍， 不利地影响多种骨骼结构的形成，包括 面部、脊柱和末端的元素。 致病基因 对于这种疾病，FGD 1编码鸟嘌呤核苷酸交换因子（GEF） 特异性激活Cdc 42，Rho（Ras同源）家族成员 p21 GTPases 通过激活Cdc 42，FGD 1刺激成纤维细胞， 形成丝状伪足，参与细胞信号传导的细胞骨架元件， 粘附和迁移。 通过Cdc 42，FGD 1还激活了 应激激活蛋白激酶（SAPK）信号级联，一种 调节细胞生长和分化。 初步研究表明， FGD 1主要在骨骼祖细胞中表达，包括椎骨 软骨前致密化、颈椎骨化和 颅面骨FGDY中受影响的骨骼 不过，数据也显示， 哺乳动物基因组中含有多种FGD 1同源物，这一结果表明， FGD 1可能是部分冗余Cdc 42激活的一个组成部分， 系统 总之，累积的数据表明FGD 1是一种组分， 一种发育调节的信号通路， 形态学和哺乳动物的骨骼发生。 本应用程序的总体目标是开发一个综合模型 为了进一步阐明FGD/Cdc 42信号级联的作用， 哺乳动物的形态发生 为此，交叉杂交和简并 PCR扩增技术将用于分离额外的FGD 1 同源物;小鼠和斑马鱼FGD cDNA克隆将用于确定 胚胎发生过程中基因表达的时空模式。 为了提供精确的细胞定位，FGD 1导向的抗体将在细胞内进行定位。 用于通过免疫细胞化学研究胚胎组织。 显微注射和 体外试验将用于研究分子生物学， 从分子上剖析FGD蛋白的功能结构域。 双杂交 和其他分子技术将用于分离和表征 FGD 1/Cdc 42信号转导途径的其他成分。 表达重组FGD 1蛋白的转基因小鼠将用于研究 FGD 1表达失调的发育后果。 将产生FGD缺陷小鼠以研究FGD缺陷对发育的影响。 FGD基因型缺失。 将产生化合物FGD缺陷型小鼠， 检查FGD同系物的功能冗余。 这些分析将 为研究FGD/Cdc 42信号的分子遗传学提供了一种手段 在哺乳动物形态发生中的转导途径。 这些分析应 导致对骨骼发生、信号 转导，和负责的多样性的一般原则， 骨骼形态