Membrane type 1 matrix metalloproteinase-associated degradation of tissue inhibitor of metalloproteinase 2 in human tumor cell lines

Tissue inhibitor of metalloproteinase 2 (TIMP-2) is required for the membra ne type 1 matrix metalloproteinase (MT1-MMP)-dependent activation of pro-MM P-2 on the cell surface. MT1-MMP-bound TIMP-2 has been shown to function as a receptor for secreted pro-MMP-2, resulting in the formation of a trimole cular complex. In the presence of uncomplexed active MT1-MMP, the prodomain of cell surface-associated MMP-2 is cleaved, and activated MMP-2 is releas ed. However, the behavior of MT1-MMP-bound TIMP-2 during MMP-2 activation i s currently unknown. In this study, I-125-labeled recombinant TIMP-2 (I-125 -rTIMP-2) was used to investigate the fate of TIMP-2 during pro-MMP-2 activ ation by HT1080 and transfected A2058 cells. HT1080 and A2058 cells transfe cted with MT1-MMP cDNA (but not vector-transfected A2058 cells) were able t o bind I-125-rTIMP-2, to activate pro-MMP-2, and to process MT1-MMP into an inactive 43-kDa form, Under these conditions, I-125-rTIMP-2 bound to the c ell surface was rapidly internalized and degraded in intracellular organell es through a bafilomycin A(1)-sensitive mechanism, and I-125-bearing low mo lecular mass fragment(s) were released in the culture medium. These differe nt processes were inhibited by hydroxamic acid-based synthetic MMP inhibito rs and rTIMP-2, but not by rTIMP-1 or cysteine, serine, or aspartic protein ase inhibitors. These results support the concept that the MT1-MMP-dependen t internalization and degradation of TIMP-2 by some tumor cells might be in volved in the regulation of pericellular proteolysis.