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.
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.
Division of Cell Biology, Kihara Institute for Biological Research, Yokohama City University, Totsuka-ku, Yokohama, 244-0813, Japan.
K
K
Shofuda
K
K
Moriyama
A
A
Nishihashi
S
S
Higashi
H
H
Mizushima
H
H
Yasumitsu
K
K
Miki
H
H
Sato
M
M
Seiki
K
K
Miyazaki
D002384:Catalysis; D002460:Cell Line; D003208:Concanavalin A / Q000494:pharmacology; D004789:Enzyme Activation; D004792:Enzyme Precursors / Q000378:metabolism; D018093:Gelatinases / Q000378:metabolism*; D006801:Humans; D053517:Matrix Metalloproteinase 15; D053513:Matrix Metalloproteinase 16; D020778:Matrix Metalloproteinase 2; D053504:Matrix Metalloproteinases, Membrane-Associated; D008666:Metalloendopeptidases / Q000145:classification / Q000378:metabolism*; D010703:Phorbol Esters / Q000494:pharmacology; D012333:RNA, Messenger / Q000378:metabolism; D019716:Tissue Inhibitor of Metalloproteinase-2 / Q000378:metabolism*; D014407:Tumor Cells, Cultured
D004792:Enzyme Precursors; C507098:MMP15 protein, human; C507102:MMP16 protein, human; D010703:Phorbol Esters; D012333:RNA, Messenger; D003208:Concanavalin A; D019716:Tissue Inhibitor of Metalloproteinase-2; D018093:Gelatinases; D053517:Matrix Metalloproteinase 15; D053513:Matrix Metalloproteinase 16; D053504:Matrix Metalloproteinases, Membrane-Associated; D008666:Metalloendopeptidases; D020778:Matrix Metalloproteinase 2