Role of Tissue Inhibitor of Metalloproteinases-2 (TIMP-2) in Regulation of Pro-Gelatinase A Activation Catalyzed by Membrane-Type Matrix Metalloproteinase-1 (MT1-MMP) in Human Cancer Cells

• Published in: Journal of biochemistry (Tokyo) 1998-08, Vol.124 (2), p.462-470
• Main Authors: Shofuda, Ken-ichi, Moriyama, Kayano, Nishihashi, Ai, Higashi, Shouichi, Mizushima, Hiroto, Yasumitsu, Hidetaro, Miki, Keizaburo, Sato, Hiroshi, Seiki, Motoharu, Miyazaki, Kaoru
• Format: Article
• Language: English
• Subjects: Catalysis; Cell Line; concanavalin A; Concanavalin A - pharmacology; Enzyme Activation; Enzyme Precursors - metabolism; gelatinase A (MMP-2); Gelatinases - metabolism; Humans; Matrix Metalloproteinase 15; Matrix Metalloproteinase 16; Matrix Metalloproteinase 2; Matrix Metalloproteinases, Membrane-Associated; membrane-type matrix metalloproteinase-1 (MT1-MMP); Metalloendopeptidases - classification; Metalloendopeptidases - metabolism; Phorbol Esters - pharmacology; phorbor ester (TPA); RNA, Messenger - metabolism; tissue inhibitor of metalloproteinase-2 (TIMP-2); Tissue Inhibitor of Metalloproteinase-2 - metabolism; Tumor Cells, Cultured
• ISSN: 0021-924X
• DOI: 10.1093/oxfordjournals.jbchem.a022136

To clarify the regulatory mechanism of pro-gelatinase A (proGelA) activation at a cellular level, expression of gelatinase A (GelA), three MT-MMPs, and TIMP-2 was examined with 11 human cancer cell lines cultured in the presence and absence of stimulants. MT1-MMP mRNA was expressed in 8 cell lines, while MT2-MMP and MT3-MMP mRNAs were expressed in fewer cell lines. The cells with high proGelA activation strongly expressed MT1-MMP mRNA but not MT2-MMP and MT3-MMP mRNAs, suggesting that MT1-MMP was responsible for the proGelA activation in the cancer cells. Treatments with concanavalin A (Con A) and a phorbor ester (TPA) enhanced the MT1-MMP expression, but only Con A stimulated the proGelA activation in many cell lines. In HT1080 fibrosarcoma cells, however, TPA also stimulated the activation. The level of TIMP-2 secreted into culture medium inversely correlated with proGelA activation. For example, 2 squamous cell carcinoma lines (HSC-3 and HSC-4) and 3 HT1080 clones, which efficiently activated proGelA, secreted little TIMP-2 into medium, whereas other cell lines and other HT1080 clones, which hardly activated proGelA, secreted TIMP-2 at high levels. When HSC-3 cells were incubated with TIMP-2 protein or transfected with TIMP-2 cDNA, the proGelA activation was strongly inhibited. These results indicated that extracellular TIMP-2 was an important negative regulator of proGelA activation. However, the level of extracellular TIMP-2 was not consistent with that of TIMP-2 mRNA in some cell lines. Other experimental results suggested that TIMP-2 might be rapidly metabolized after binding to MT1-MMP, and Con A treatment might stabilize the complex of TIMP-2 and MT1-MMP on cell membranes.