Matrix Metalloproteinase 9 Regulates Tumor Cell Invasion via Cleavage of Protease Nexin-1

Matrix Metalloproteinase 9 (MMP-9) expression is known to enhance the invasion and metastasis of tumor cells. In previous work based on a proteomic screen, we identified the serpin Protease nexin-1 (PN-1) as a potential target of MMP-9. Here we demonstrate that PN-1 is a substrate for MMP-9 and establish a link between PN-1 degradation by MMP-9 and regulation of invasion. PN-1 levels increased in prostate carcinoma cells after downregulation of MMP-9 and in tissues of MMP-9 deficient mice, consistent with MMP-9 acting to degrade PN-1. We identified three MMP-9 cleavage sites in PN-1 and demonstrated that mutations in those sites made PN-1 more resistant to MMP-9. Urokinase plasminogen activator (uPA) is inhibited by PN-1. MMP-9 augmented uPA activity in the medium of PC3-ML cells by degrading PN-1. Prostate cancer cells, overexpressing PN-1 or treated with MMP-9 shRNA, had reduced cell invasion in matrigel. PN-1 siRNA restored uPA activity and the invasive capacity. PN-1 mutated in the serpin inhibitory domain, the reactive centre loop (RCL), failed to inhibit uPA and failed to reduce matrigel invasion. Taken together, this study demonstrates a novel molecular pathway in which MMP-9 regulates uPA activity and tumor cell invasion through cleavage of PN-1.