A receptor for tPA was previously described on human umbilical vein endothelial cells (Annual Report 1986, human epidermoid carcinoma cells (A- 431), and human osteosarcoma cells (HOS) (Annual Report 1989). The present work addressed the characterization of this receptor. Crosslinking studies with these cell lines indicated that the receptor was in the molecular weight range of 54,000 as shown by HPLC and SDS-PAGE analyses. Isolation of the putative receptor was attempted by passing cell lysates over a t-PA conjugated affinity column. Several t-PA binding proteins in the molecular weight range of 54,000 were isolated. Ligand blotting with affinity purified cell lysates similarly revealed reactive bands in the 54,000 molecular weight range. Several of the contaminating proteins were identified by immunoblotting and included tubulin beta chain, plasminogen activator (scu-PA). The identity of tubulin beta chain was confirmed by N- terminal sequence analysis. Efforts to identify a specific receptor protein are continuing. In related studies the ability of cells to bind t-PA was shown not to be related to the synthesis of mRNA of any of the fibrinolytic proteins normally produced by various cell types (t-PA, u-PA, or PAI-1). However, enhanced binding has been repeatedly demonstrated in the presence of exogenously added suc-PA. Current work focuses on the interaction of t-PA with scu-PA.
