Prieto A L, Andersson-Fisone C, Crossin K L
Rockefeller University, New York, NY 10021.
J Cell Biol. 1992 Nov;119(3):663-78. doi: 10.1083/jcb.119.3.663.
The extracellular matrix molecule cytotactin is a multidomain protein that plays a role in cell migration, proliferation, and differentiation during development. To analyze the structure-function relationships of the different domains of this glycoprotein, we have prepared a series of fusion constructs in bacterial expression vectors. Results obtained using a number of adhesion assays suggest that at least four independent cell binding regions are distributed among the various cytotactin domains. Two of these are adhesive; two others appear to be counteradhesive in that they inhibit cell attachment to otherwise favorable substrates. The adhesive regions were mapped to the fibronectin type III repeats II-VI and the fibrinogen domain. The morphology of the cells plated onto these adhesive fragments differed; the cells spread on the fibronectin type III repeats as they do on fibronectin, but remained round on the fibrinogen domain. The counteradhesive properties of the molecule were mapped to the EGF-like repeats and the last two fibronectin type III repeats, VII-VIII. The latter region also contained a cell attachment activity that was observed only after proteolysis of the cells. Several cell types were used in these analyses, including fibroblasts, neurons, and glia, all of which are known to bind to cytotactin. The different domains exert their effects in a concentration-dependent manner and can be inhibited by an excess of the soluble molecule, consistent with the hypothesis that the observed properties are mediated by specific receptors. Moreover, it appears that some of these receptors are restricted to particular cell types. For example, glial cells bound better than neurons to the fibrinogen domain and fibroblasts bound better than glia and neurons to the EGF fragment. These results provide a basis for understanding the multiple activities of cytotactin and a framework for isolating different receptors that mediate the various cellular responses to this molecule.
细胞外基质分子细胞趋触蛋白是一种多结构域蛋白，在发育过程中的细胞迁移、增殖和分化中发挥作用。为了分析这种糖蛋白不同结构域的结构 - 功能关系，我们在细菌表达载体中制备了一系列融合构建体。使用多种黏附试验获得的结果表明，至少四个独立的细胞结合区域分布在细胞趋触蛋白的各个结构域中。其中两个区域具有黏附性；另外两个区域似乎具有抗黏附性，因为它们会抑制细胞附着在原本有利的底物上。黏附区域被定位到纤连蛋白III型重复序列II - VI和纤维蛋白原结构域。接种到这些黏附片段上的细胞形态不同；细胞在纤连蛋白III型重复序列上的铺展方式与在纤连蛋白上相同，但在纤维蛋白原结构域上仍保持圆形。该分子的抗黏附特性被定位到表皮生长因子样重复序列和最后两个纤连蛋白III型重复序列，VII - VIII。后一个区域还含有一种细胞附着活性，这种活性只有在细胞经过蛋白酶解后才能观察到。这些分析使用了几种细胞类型，包括成纤维细胞、神经元和神经胶质细胞，所有这些细胞都已知会与细胞趋触蛋白结合。不同的结构域以浓度依赖的方式发挥作用，并且可以被过量的可溶性分子抑制，这与观察到的特性是由特定受体介导的假设一致。此外，似乎其中一些受体仅限于特定的细胞类型。例如，神经胶质细胞比神经元更好地结合纤维蛋白原结构域，而成纤维细胞比神经胶质细胞和神经元更好地结合表皮生长因子片段。这些结果为理解细胞趋触蛋白的多种活性提供了基础，并为分离介导细胞对该分子各种反应的不同受体提供了框架。
