STRUCTURE/FUNCTION AND SIGNAL TRANSDUCTION PATHWAYS OF TGF BETA

TGF Beta 的结构/功能和信号转导途径

• 批准号: 6160869
• 负责人: A B ROBERTS
• 金额: --
• 依托单位: DIVISION OF BASIC SCIENCES - NCI
• 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/6160869
• 关键词: biological signal transduction; genetically modified animals; growth factor receptors; laboratory mouse; protein isoforms; protein kinase; protein structure function; receptor binding; receptor coupling; transcription factor; transforming growth factors; tumor suppressor genes; yeast two hybrid system

The TGF-b family of ligands signal through a unique heteromeric receptor complex distinguished by its serine-threonine kinase activity. One signal transduction pathway from these receptors involves a novel family of proteins termed Smads, which are related to Drosophila Mad, a cytoplasmic mediator of signals from a TGF-b family ligand, dpp. We have demonstrated that one of these, Smad4, identical to a candidate tumor suppressor for pancreatic cancer, DPC4, can restore TGF-b signal transduction pathways in tumor cells null for Smad4. By constructing chimeric and mutated forms of Smad4, we have shown that the ligand- dependency resides in its unique proline-rich linker and N-terminal domain inserts. Ongoing studies are aimed at characterization of the functional activity associated with these domains. Other studies are focused on characterization of two novel clones we have isolated from a yeast two-hybrid system using Smad1 as bait. The proteins show no identifiable enzymatic motifs or homology to existing proteins. They suggest additional complexity to these signal transduction pathways in that they inhibit Smad-induced reporter gene activity. We are currently characterizing the interaction patterns and mechanisms of action of these proteins. In other studies, we have demonstrated that Smads may mediate cross-talk between serine-threonine kinase receptors and certain tyrosine kinase receptors, in that both HGF and EGF can phosphorylate Smad1 and Smad2 in cells lacking TGF-b receptors. HGF can also stimulate nuclear translocation of Smad2, suggesting that the overlapping activities of TGF-b and HGF in activation of certain target genes such as tissue inhibitor of metalloproteinases (TIMP) and plasminogen activator inhibitor-1 (PAI-1) are mediated by a Smad-dependent pathway. To delineate more fully the extent of Smad-mediated pathways in transduction of signals from TGF-b family ligands, we have generated mice null for Smad1 and plan to create transgenic mice expressing dominant Smad1 and 2 under control of tissue-specific promoters. The Smad1 null mice die between days 8 and 9 of embryogenesis, demonstrating a critical signaling role for this molecule in early development. We plan to backcross transgenic mice expressing dominant negative Smads with mice heterozygous for Smad1 and for Smads 2, 3, and 4 (obtained collaboratively from Chuxia Deng) to enhance the phenotype of compromised Smad activity.

TGF-β配体家族通过一种独特的异聚体受体传递信号 复合物的特征在于其丝氨酸-苏氨酸激酶活性。一 这些受体的信号转导途径涉及一个新的家族 一种名为Smads的蛋白质，与果蝇Mad有关， 来自TGF-β家族配体的信号的细胞质介质，dpp。我们有 证明了其中一个Smad4与候选肿瘤相同， 胰腺癌抑制因子DPC4可以恢复TGF-β信号 肿瘤细胞中Smad4无效的转导途径。通过构造 嵌合和突变形式的Smad4，我们已经表明，配体- 依赖性存在于其独特的富含脯氨酸的接头和N-末端 域插入。正在进行的研究旨在表征 与这些领域相关的功能活动。其他研究 重点是表征我们分离出的两个新克隆， 以Smad1为诱饵的酵母双杂交系统。蛋白质显示没有 可识别的酶基序或与现有蛋白质的同源性。他们 这表明这些信号转导途径在 它们抑制Smad诱导的报告基因活性。我们目前正在 描述的相互作用模式和作用机制， 这些蛋白质。在其他研究中，我们已经证明，Smads可能 介导丝氨酸-苏氨酸激酶受体与某些 酪氨酸激酶受体，因为HGF和EGF都可以磷酸化 Smad1和Smad2在缺乏TGF-β受体的细胞中的表达。HGF还可以刺激 Smad2的核转位，这表明， TGF-β和HGF在某些靶基因的激活中的活性， 作为金属蛋白酶组织抑制剂（TIMP）和纤溶酶原 活化剂抑制剂-1（PAI-1）的作用是由Smad依赖性途径介导的。 为了更全面地描述Smad介导的通路在 从TGF-β家族配体的信号转导，我们已经产生 Smad1无效小鼠，并计划创建表达 在组织特异性启动子控制下的显性Smad1和2。的 Smad1基因敲除小鼠在胚胎发生的第8 - 9天死亡，表明 这是这种分子在早期发育中的关键信号作用。我们 计划回交表达显性阴性Smads的转基因小鼠 用Smad 1和Smads 2、3和4杂合子小鼠（获得 来自Chuxia Deng的合作）以增强 Smad活动受损