Structural and Functional Analysis of New Type Extracellular Matrix Receptor syndecan-2

小鼠Lewis肺癌来源的低转移P29克隆的肿瘤发生依赖于富含纤维连接蛋白的基质基质，而高转移LM66-H11克隆的生长依赖于基底膜。在体外黏附于纤维连接蛋白底物上，P29细胞形成应力纤维，而LM66-H11细胞形成皮质作用结构。P29细胞的表型与整合素α-5、β-1和Syndrecan-2的表达密切相关，这些整合素的侧链与COOH端的肝素结合区具有特异性的亲和力。LM66-H11细胞的整合素表达水平与P29细胞相同，而Syndecan-2的表达水平明显低于P29细胞。体外用Syndecan-2反义寡核苷酸处理P29细胞，选择性抑制其生物合成，其表型与LM66-H11细胞无明显差别。P29细胞的表型是通过与纤维连接蛋白重组融合多肽(CH-271)的粘附来复制的，该融合多肽包括RGD细胞结合(C-274)和C端肝素结合(H271)结构域，而LM66-H11细胞的表型仅由C-274多肽诱导，这表明整合素α5β1和Syndecan-2信号对多肽是必需的，该多肽可以被大量的H-271多肽或针对Syndecan-2或Hean的抗体所取代。此外，当接种到由C-274多肽和碱性成纤维细胞生长因子组成的融合多肽时，即使是LM66-H11细胞也形成了应力纤维。这些发现表明，有必要将这两个受体聚集在一起，以发出组织肌动蛋白细胞骨架的信号。

Mouse Lewis lung carcinoma-derived low metastatic P29 clone exhibits tumorigenesis dependent on the fibronectin-rich stromal matrix, whereas the growth of highly metastatic LM66-H11 clone depends on the basement membranes. On adhesion to the fibronectin substratum in vitro, P29 cells show stress fiber formation, whereas LM66-H11 cells form cortex action structure. The phenotype of P29 cells is closely correlated to the expression of integrin α5β1 and syndrecan-2 having heparan sulfate side chains with specific affinity to COOH-terminal heparin-binding domain. LM66-H11 cells express the integrin at the same level to that of P29 cells, but syndecan-2 at a significantly lower level. On treatment of P29 cells in vitro with antisense oligonucleotide for syndecan-2, causing selective inhibition of its biosynthesis, the cells turn out to be indistinguishable from LM66-H11 cells as regards the phenotype. The phenotype of P29 cells is reproduced by adhesion to fibronectin recombinant fusion polypeptide (CH-271) comprising RGD cell-binding (C-274) and C-terminal heparin-binding (H271) comprising RGD cell-binding (C-274) and C-terminal heparin-binding (H271) domains whereas the phenotype of LM66-H11 cells in induced only by C-274 polypeptide, indicating that signaling through integrin α5β1 and syndecan-2 is essential for the polypeptide can be replaced by a large amount of H-271 polypeptide, or antibodies specific to syndecan-2 or heparan sulfate. Furthermore, when inoculated on a fusion polypeptide comprising C-274 polypeptide and basic FGF with strong affinity to syndecan-2 ectodomain, even LM66-H11 cells form stress fibers. These findings indicate the necessity of clustering of the two receptors for signaling for the organization of the actin cytoskeletons.