Thrombin has been shown to activate tumor-cell adhesion to platelets, fibronectin and von Willebrand factor 2- to 3-fold in vitro, and enhance metastasis 10- to 156-fold in vivo. We therefore elected to determine whether thrombin binds to tumor cells and whether thrombin-treated tumor cells enhance their adhesion to endothelial cells, the first barrier to tumor invasion and metastasis. Thrombin-treated human and hamster melanoma cells (SK-Mel-28 and HM-29) enhanced their adhesion to bovine aortic endothelial cells 2.1- to 2.3-fold, respectively. Similar results were obtained with bovine capillary endothelial cells. Thrombin activation of tumor cells was rapid, reaching its peak 15 min after thrombin activation; and transient, declining to baseline levels by 60 min. 125I-thrombin bound to both SK-Mel-28 and HM-29 cells in a saturation-dependent manner, was inhibitable by unlabelled thrombin, and could be 90% washed away with buffer following 30 min of incubation. Electron microscopy of tumor cells bound to fibronectin-coated millipore filters revealed adhesion of naive as well as thrombin-treated tumor cells to endothelial cells and subendothelial matrix between endothelial cells. Neither mode of adhesion was preferentially enhanced by thrombin-treated tumor cells. Both naive and thrombin-treated SK-Mel-28 cells had the adhesive ligand integrin receptors: alpha 3 beta l (fibronectin, laminin, collagen); alpha 5 beta l (fibronectin); alpha v beta x (vitronectin). Receptors for the beta 2 integrin family (LFA-I and Mac-I) were not found, nor were receptors of the beta 3 integrin family, GPIIIa. The receptor ligands fibronectin and vitronectin were present. None of the above receptors or ligands increased their density or appeared de novo after thrombin stimulation. Thus, 2 melanoma cell lines have thrombin receptors which, when occupied, lead to enhanced adhesion of tumor cells to endothelium and subendothelial matrix.