| Generation of plasmin is central to cell movement during wound healing, tumor metastasis, and inflammation, and is accelerated when plasminogen binds to cell surface receptors bearing lysine at their carboxy-terminus. The identity of the plasminogen receptor(s) responsible for regulating plasmin generation at the cell surface is currently unclear. In this study we found that the candidate plasminogen receptor, annexin A2 heterotetramer, accelerates the generation of plasmin by tissue-type plasminogen activator (tPA) and urokinase plasminogen activator (uPA), and that this acceleration is dependent upon lysine residues at the carboxy-terminus of the S100A10 subunit of the heterotetramer. Physiological concentrations of plasma carboxypeptidases were capable of abrogating the effect on plasmin generation, by removal of both the ultimate and penultimate lysine residues from S100A10, suggesting a possible mechanism of regulating plasmin formation in vivo. It was found that S100A10 expression was induced in the monocytoid cell lines, THP-1 and U937, by high cell culture density, which was dependent upon de novo protein synthesis. Increased cell surface expression of S100A10 was induced upon differentiation of THP-1 cells by exposure to phorbol 12-myristate 13-acetate, interferon gamma, fibronectin, and 1alpha, 25-dihydroxyvitamin D3 and correlated with an increase in cellular plasminogen binding. Annexin A2 heterotetramer localized to discrete regions of the surface of a subpopulation of cells after polarization. This distribution overlapped to a great degree with that of the uPA receptor, which is known to redistribute to the leading edge of polarized cells. Importantly, plasminogen was also found to localize to these discrete membrane regions and also largely colocalized with S100A10. Specific downregulation of S100A10 expression by small interfering RNA led to a decrease in extracellular levels of both S100A10 and annexin A2, as well as a decrease in the rate of plasminogen activation on the surface of THP-1 cells. In addition, invasion of both THP-1 cells and K562 cells through physiological extracellular matrix was inhibited by siRNA-mediated reduction of S100A10 expression. Thus the current work establishes that S100A10 is a regulated cell surface protein that is directly involved in cellular plasmin production and contributes to the invasiveness of monocytoid cells. |
