Expression and colocalization of cytokeratin 1 and urokinase plasminogen activator receptor on endothelial cells

The cellular localization of human cytokeratin 1 (CK1), urokinase plasminog en activator receptor (UPAR), and gClqR, high-molecular-weight kininogen (H K)-binding proteins on endothelial cells, was determined. CK1 was found on the external membrane of nonpermeabilized endothelial cells by immunoperoxi dase staining, immunofluorescence, and transmission electron microscopy usi ng immunogold. Human umbilical vein endothelial cells (HUVECs) had 7.2 +/- 0.2 x 10(4) specific CK1 membrane sites/cell by I-125-F(ab')(2) anti-CK1 an tibody binding. Flow cytometry studies confirmed the presence of CK1, uPAR, and gClqR on HUVECs. On laser scanning confocal microscopy and transmissio n electron microscopy, CK1 and uPAR, but not gClqR, colocalized on the cell surface of HUVECs. The HUVEC surface distribution of these proteins was di stinctly different from that for von Willebrand factor. In competitive inhi bition experiments, anti-CK1, anti-uPAR, or anti-gC1qR blocked both biotin- HK binding and prekallikrein (PK) activation on HUVECs with an inhibitory c oncentration of 50% (IC50) of 300 to 350 nM, 50 to 60 nM, or 35 to 100 nM, respectively. Also, antibodies to uPAR and gClqR each inhibited 86% of kall ikrein-mediated, P-chain urokinase plasminogen activation, whereas antibodi es to CK1 only inhibited 24% of plasminogen activation. On HUVECs, CK1 and uPAR, but not gClqR, colocalized to be a multiprotein receptor complex for HK binding, PK activation, and 2-chain urokinase plasminogen activation. (B lood. 2001;97:2342-2350) (C) 2001 by The American Society of Hematology.