STRUCTURE/FUNCTION AND SIGNAL TRANSDUCTION PATHWAYS OF TGF BETA

转化生长因子-β配体家族通过一种独特的异构体受体传递信号 以其丝氨酸-苏氨酸激酶活性而区别的复合体。一 这些受体的信号转导途径涉及一个新的家族 在与果蝇Mad相关的名为Smads的蛋白质中， 转化生长因子-b家族配体DPP信号的细胞质介体。我们有 证明了其中一种，Smad4，与候选肿瘤相同 胰腺癌抑制因子DPC4可恢复转化生长因子-b信号 肿瘤细胞中Smad4的转导通路为空。通过构建 Smad4的嵌合和突变形式，我们已经证明了配体- 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描述 与这些区域相关的功能活动。其他研究包括 重点研究了我们从其中分离的两个新克隆的特征 以Smad1为诱饵的酵母双杂交系统。这些蛋白质显示没有 可识别的酶基序或与现有蛋白质的同源性。他们 提示这些信号转导通路的额外复杂性 它们抑制Smad诱导的报告基因活性。我们目前正在 描述相互作用的模式和作用机制 这些蛋白质。在其他研究中，我们已经证明了SMADS可能 介导丝氨酸苏氨酸激酶受体和某些 酪氨酸激酶受体，因为HGF和EGF都可以磷酸化 Smad1和Smad2在缺乏转化生长因子-b受体的细胞中。肝细胞生长因子也可以刺激 Smad2的核易位，提示重叠 转化生长因子-β和肝细胞生长因子在某些靶基因激活中的活性 作为金属蛋白酶组织抑制物(TIMP)和纤溶酶原 激活物抑制因子-1(PAI-1)是由Smad依赖的途径介导的。 为了更全面地描述Smad介导的通路在 转化生长因子-β家族配体的信号转导，我们已经产生了 小鼠没有Smad1基因，并计划创建表达Smad1的转基因小鼠 组织特异性启动子控制下的优势Smad1和Smad2。这个 Smad1缺失的小鼠在胚胎发育的第8天到第9天死亡，证明 这种分子在早期发育中起着关键的信号作用。我们 计划回交表达显性负Smads的转基因小鼠 具有Smad1和Smad 2、3和4杂合子的小鼠(获得 出自邓楚霞)，以增强表型 泄露了Smad的活动。

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.