BIOLOGY AND RECEPTOR SIGNALLING OF TRANSFORMING GROWTH FACTOR-BETA

在哺乳动物中表达的转化生长因子-β的三种亚型在功能上是 在大多数体外测试系统中可互换，但它们具有独特的 某些细胞上的活动。确定特定的功能区域 转化生长因子-β，以及确定关键的受体成分 转化生长因子-β作用，我们正在利用结肠癌细胞，其中 转化生长因子-β1抑制生长的活性是它的几百倍 比转化生长因子-β2更有效。定义的特定区域 转化生长因子-β分子负责这些活动的差异，我们 建立了重组蛋白的表达和纯化系统 转化生长因子-β基因被设计为具有突变或替换 不同亚型的同源区域，基于三维 转化生长因子-β的结构。利用结直肠癌细胞显示 对于转化生长因子-β1和3的选择性，我们已经确定了另一个区域 可能参与的分子(氨基酸69-73) 异构体特异性受体结合。我们还演示了，使用 测定体外与可溶性转化生长因子-βII和 III受体和转化生长因子-β潜伏期相关蛋白(LAP)， 与这些蛋白质相互作用的特定配基区域。在……里面 这个问题的一个相关方面，我们试图描述 转化生长因子-β受体在结肠癌细胞上的表达 确定可能的下游信令中间体。这些研究 在识别受体的特定元件时都很重要 介导异构体特异性效应的结合系统，并在识别 假定的信号中间体，其缺失或突变 会导致转化生长因子-β反应性的丧失。现建议： 这样的中间体将具有肿瘤抑制因子样的活性。

The three isoforms of TGF-beta expressed in mammals are functionally interchangeable in most in vitro assay systems, but they have distinctive activities on certain cells. To identify specific functional regions of the TGF-betas as well as to identify receptor components critical for TGF-beta action, we are utilizing colon carcinoma cells where the activities of TGF-beta1 on inhibition of growth are several hundred-fold more potent than that of TGF-beta2. To define specific regions of the TGF-beta molecule responsible for these differences in activities, we developed a system for expression and purification of recombinant TGF-betas engineered to have either mutations or substitutions of homologous regions of different isoforms, based on the 3-dimensional structure of TGF-beta. Using colorectal carcinoma cells which show selectivity for TGF-beta 1 and 3, we have identified another region of the molecule (amino acids 69-73) which may be involved in isoform-specific receptor binding. We have also demonstrated, using assays which measure in vitro binding to the soluble TGF-beta type II and III receptors and to the TGF-beta latency-associated protein (LAP), specific regions of the ligands which interact with these proteins. In a related aspect of this problem, we are attempting to characterize the expression of TGF-beta receptors on the colon carcinoma cells and to identify possible down-stream signalling intermediates. These studies are important both in identifying the specific elements of the receptor binding system which mediate isoform-specific effects, and in identifying putative signalling intermediates, the deletion or mutation of which would result in loss of TGF-beta responsiveness. It is proposed that such intermediates will have tumor-suppressor-like activity.