CHARACTERIZATION OF A NOVEL, MEMBRANE-BOUND, 80-KDA MATRIX-DEGRADING PROTEASE FROM HUMAN BREAST-CANCER CELLS - MONOCLONAL-ANTIBODY PRODUCTION, ISOLATION, AND LOCALIZATION

A major, apparently novel extracellular matrix-degrading protease was previously identified and partially isolated from hormone-dependent bu t not from hormone-independent human breast cancer cells (Shi, Y. E., Torri, J., Yieh, L., Wellstein, A., Lippman, M. E., and Dickson, R. B. (1993) Cancer Res. 53, 1409-1415). Although initially the 80-kDa prot ease was identified from breast cancer cell-conditioned medium, immuno fluorescence staining of breast cancer cells with anti-80-kDa protease monoclonal antibody 21-9 showed that in addition to its detection in intracellular compartments, the protease was uniformly localized aroun d periphery of the cells with more intensive staining on the pseudopod ia and membrane ruffles. A surface biotinylation technique confirmed t he plasma membrane localization of the protease. In addition, the 80-k Da protease could not be washed from the membrane fraction of homogeni zed breast cancer cells with high concentrations of salts or with EDTA . The 80-kDa protease may noncovalently associate with other protein(s ) to form complexes, the 95- and 110-kDa proteases. Both complexes sho wed gelatinolytic activity and bore the epitopes recognized by monoclo nal antibody 21-9. Furthermore, both complexes could be converted to 8 0-kDa forms by boiling in SDS in the absence of reducing agents. Expre ssion of this novel, integral membrane gelatinase could allow breast c ancer cells an alternative to other previously described matrix-degrad ing enzymes for degradation of the extracellular matrix in close proxi mity to their surfaces.