Cell surface association of extracellular matrix (ECM)-degrading enzymes has been suggested to facilitate proteolysis of ECM in areas of cell-matrix contacts and to be crucial for the process of tumor cell invasion. Matrix metalloproteinase-9 (MMP-9) is a member of the MMP family of endopeptidases that has been shown to play a critical role in hydrolysis of ECM components and has been localized on the surface of tumor cells. However, the nature of the cell surface association of MMP-9 is unknown. Here, we report the cell surface association of MMP-9 in human breast epithelial MCF10A cells treated with 12-O-tetradecanoylphorbol-13-acetate (TPA). Surface biotinylation and immunoprecipitation with anti-MMP-9 antibodies revealed the presence of two MMP-9 forms (Mr 92,000 and 85,000) on the surface of TPA-treated MCF10A cells, whereas in the media, only the Mr 92,000 form was detected, mostly in complex with TIMP-1, a specific MMP-9 inhibitor. The MMP-9 forms were also found in purified plasma membranes of TPA-treated cells. In contrast, the plasma membranes contained little or no TIMP-1. The surface-bound MMP-9 forms were recognized by an antibody to the NH2-terminal prodomain, indicating that both represent latent enzymes. Pulse-chase analysis and endoglycosidase H digestion of surface-biotinylated MMP-9 forms demonstrated that the Mr 85,000 species was endoglycosidase H sensitive, suggesting targeting of the precursor form of MMP-9 to the cell surface. These studies demonstrate a specific cell surface association of MMP-9 in response to TPA that may help to localize TIMP-1-free enzyme on the surface of breast epithelial cells.

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Supported by Grant CA61986 from the National Cancer Institute, NIH and by Department of Defense Grant DAMD17-94-J-4356 (to R. F.).

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©1997 American Association for Cancer Research.
1997
American Association for Cancer Research, Inc.