Plasma Membrane-bound Tissue Inhibitor of Metalloproteinases (TIMP)-2 Specifically Inhibits Matrix Metalloproteinase 2 (Gelatinase A) Activated on the Cell Surface*
- From the ‡Department of Biochemistry and Molecular Biology, The University of Kansas Medical Center, Kansas City, Kansas 66160-7421, the ¶Department of Biochemistry, Tokyo University of Pharmacy and Life Science, School of Pharmacy, Horinouchi, Hachioji, Tokyo 192-0392, the‖Biopharmaceutical Department, Fuji Chemical Industries, Ltd., Takaoka, Toyama 933-8511, and **Biological Research Laboratories, Sankyo Co., Ltd., Shinagawa-ku, Tokyo 140-8710, Japan
Abstract
The cell-surface activation of pro-matrix metalloproteinase 2 (pro-MMP-2) is considered to be critical for cell migration and invasion. Treatment of human uterine cervical fibroblasts with concanavalin A activates pro-MMP-2 on the cell surface by converting it to the 65-kDa form with a minor form of 45 kDa. However, the 65-kDa MMP-2 was inactivated by tissue inhibitor of metalloproteinases (TIMP)-2 that was bound to the plasma membrane upon concanavalin A treatment. TIMP-2 binds to the plasma membrane through its N-terminal domain by two different modes of interaction as follows: one is sensitive to a hydroxamate (HXM) inhibitor of MMPs and the other is HXM-insensitive. TIMP-2 bound to the membrane in a HXM-insensitive manner, comprising about 40–50% of TIMP-2 on the membrane, is the inhibitor of the cell surface-activated MMP-2. It, however, does not inhibit MMP-3, MMP-9, and the 45-kDa MMP-2 lacking the C-terminal domain. The inhibition of the 65-kDa MMP-2 by TIMP-2 is initiated by the interaction of their C-terminal domains. Subsequently, the MMP-2·TIMP-2 complex is released from the membrane, and the activity of MMP-2 is blocked by TIMP-2. In the presence of collagen types I, II, III, V, or gelatin, the rate of inhibition of the 65-kDa MMP-2 by the membrane-bound TIMP-2 decreased considerably. These results suggest that the pericellular activity of MMP-2 is tightly regulated by membrane-bound TIMP-2 and surrounding extracellular matrix components.
Footnotes
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↵* This work was supported by National Institutes of Health Grants AR 39189 and AR 40994.The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
Dedicated to Dr. J. Frederick Woessner, Jr., on the occasion of his 70th birthday.
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↵§ Present address: Dept. of Cancer Cell Research, Institute of Medical Science, University of Tokyo, 4-6-1 Shirokanedai, Minato-ku, Tokyo 108-8639, Japan.
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↵‡ To whom all correspondence should be addressed: Hideaki Nagase, Dept. of Biochemistry and Molecular Biology, University of Kansas Medical Center, 3901 Rainbow Blvd., Kansas City, KS 66160-7421. Tel.: 913-588-7079; Fax: 913-588-7111; E-mail: hnagase{at}kumc.edu.
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↵2 Y. Itoh, and H. Nagase, unpublished observations.
- Abbreviations:
- MMP
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matrix metalloproteinase
- APMA
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4-aminophenylmercuric acetate
- TIMP
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tissue inhibitor of metalloproteinases
- ConA
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concanavalin A
- α2M
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α2-macroglobulin
- HXM
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hydroxamic acid MMP inhibitor
- E-64
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trans-epoxysuccinyl-l-leucylamido(4-guanidino)-butane
- PAGE
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polyacrylamide gel electrophoresis
- DMEM
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Dulbecco’s modified Eagle’s medium
- HBSS
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Hanks’ balanced salt solution
- Mca
-
(7-methoxycoumarin-4-yl)acetyl
- Dpa
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3-(2,4-dini- trophenyl)-l-2,3-diaminopropionyl.
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- Received April 24, 1998.
- Revision received July 6, 1998.
- The American Society for Biochemistry and Molecular Biology, Inc.
