Adams J C
Medical Research Council Laboratory for Molecular Cell Biology, University College London, United Kingdom.
Mol Biol Cell. 1997 Nov;8(11):2345-63. doi: 10.1091/mbc.8.11.2345.
Cell adhesion to thrombospondin-1 (TSP-1) correlates with assembly of cell-substratum contact structures that contain fascin microspikes. In this analysis, cell-matrix requirements for assembly of fascin microspikes were examined in detail. In six cell lines, cell spreading on a TSP-1 substratum correlated with expression of fascin protein and formation of fascin microspikes. Microspikes were not formed by H9c2 cells adherent on fibronectin, vitronectin, collagen IV, or platelet factor 4. However, both fascin microspikes and focal contacts were assembled by cells adherent on laminin-1. Using mixed substrata containing different proportions of TSP-1, and fibronectin, fascin microspike formation by H9c2 and C2C12 cells was found to be reduced on substrata containing 25% fibronectin and abolished on substrata containing 75% fibronectin. Adhesion to intermediate mixtures of TSP-1 and fibronectin resulted in coassembly of fascin microspikes and focal contacts, colocalization of fascin with actin stress fiber bundles and altered distributions of beta 1 integrins, cortical alpha-actinin, and tropomyosin. In cells adherent on 50% TSP-1:50% fibronectin, GRGDSP peptide treatment decreased focal contact assembly and altered cytoskeletal organization but did not inhibit microspike assembly. Treatment with chondroitin sulfate A or p-nitrophenol beta-D-xylopyranoside decreased microspike formation and modified cytoskeletal organization but did not inhibit focal contact formation. In polarized migratory and postmitotic C2C12 cells, fascin microspikes and ruffles were localized at leading edges and TSP matrix deposition was also concentrated in this region. Depletion of matrix TSP by heparin treatment correlated with decreased microspike formation and cell motility. Thus, the balance of adhesive receptors ligated at the cell surface during initial cell-matrix attachment serves to regulate the type of substratum adhesion contact assembled and subsequent cytoskeletal organization. A role for fascin microspikes in cell motile behavior is indicated.
细胞与血小板反应蛋白-1（TSP-1）的黏附与包含丝束蛋白微刺的细胞-基质接触结构的组装相关。在本分析中，详细研究了丝束蛋白微刺组装所需的细胞-基质条件。在六种细胞系中，细胞在TSP-1基质上的铺展与丝束蛋白的表达及丝束蛋白微刺的形成相关。贴附于纤连蛋白、玻连蛋白、IV型胶原或血小板因子4上的H9c2细胞不会形成微刺。然而，贴附于层粘连蛋白-1上的细胞会组装丝束蛋白微刺和黏着斑。使用含有不同比例TSP-1和纤连蛋白的混合基质，发现H9c2和C2C12细胞在含有25%纤连蛋白的基质上丝束蛋白微刺形成减少，而在含有75%纤连蛋白的基质上则完全不形成。与TSP-1和纤连蛋白的中间混合物黏附会导致丝束蛋白微刺和黏着斑共同组装，丝束蛋白与肌动蛋白应力纤维束共定位，并改变β1整合素、皮层α-辅肌动蛋白和原肌球蛋白的分布。在贴附于50% TSP-1:50%纤连蛋白的细胞中，GRGDSP肽处理会减少黏着斑组装并改变细胞骨架组织，但不抑制微刺组装。硫酸软骨素A或对硝基苯酚β-D-吡喃木糖苷处理会减少微刺形成并改变细胞骨架组织，但不抑制黏着斑形成。在极化迁移和有丝分裂后的C2C12细胞中，丝束蛋白微刺和褶皱位于前缘，TSP基质沉积也集中在该区域。肝素处理使基质TSP耗竭与微刺形成减少和细胞运动性降低相关。因此，在初始细胞-基质黏附过程中细胞表面连接的黏附受体平衡有助于调节组装的基质黏附接触类型及随后的细胞骨架组织。这表明丝束蛋白微刺在细胞运动行为中起作用。
