1993 Fiscal Year Final Research Report Summary
Molecular mechanism of fibronectin-induced chemotactic migration
| Project/Area Number |
04680173
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| Research Category |
Grant-in-Aid for General Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Research Field |
物質生物化学
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| Research Institution | Faculty of Pharmaceutical Sciences, Science University of Tokyo |
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Principal Investigator |
FUKAI Fumio Science Univ.of Tokyo, 薬学部, 講師 (90124487)
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| Co-Investigator(Kenkyū-buntansha) |
KATAYAMA Takashi Science Univ.of Tokyo, 薬学部, 教授 (00013897)
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| Project Period (FY) |
1992 – 1993
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| Keywords | fibronectin / fibrofnectin fragment / cell migration / protease / matrix metalloproteinase / cell differentiation / receptor / calpain |
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| Research Abstract |
We have shown that proteolytic degradation of FN yields biological activities not present in intact FN.We found that 21K Fib 2 fragment has a chemotactic activity at extremely low concentrations that intact FN hardly show the activity. Adipocyte differentiation of ST-13 preadipocytes is inhibited by intact FN, but the 24K fragment originated fromtheamino-terminal fibrin-binding domain of FN molecule dramatically stimulated the differentiation. The mode of proteolytic degradation of intact FN may be an important determinant for activation of FN function. First, we examined effects of inflammatory proteases (plasmin, neutrophil cathepsin G and elastase) and matrix metalloproteinases (stromelysin, matrilysin, gelatinase A, and interstitial collagenase) on the release of active FN fragments. Gelatinase A was found to be the most active in generating chemotactic Fib 2 fragments. Differentiation stimuratory Fib 1 fragments were effectively released by neutrophil elastase. In addition to such
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indirect action of proteases on the chemotactic migration, some intracellular Ca^<2+> dependentcysteine proteases were shown to involve directly in the chemotactic migration in response to FN.
Second, a cell surface receptor protein for the 21K Fib 2 fragment was tried to isolate by an affinity column using 21K Fib 2 fragment-fixed gel. Washing the column with GRGDSP peptide, BSA solution, and then the Fib 2 fragment-depleted FN fragments mixture, after applying the ^<125>I-labed NIH-L13 cell extract, enabled to remove exclusively the proteins binding nonspecifically to the affinity gel. As a result, one sharp radioactive peak could be eluted with the 21K Fib 2 fragment, in which three protein bands with Mr.of 30kDa, 60kDa, and >200kDa were detected by aotoradiography. Cross-linking of the 21K Fib 2 fragment and cell surface receptor with bireactive agent DSS also showed that the Fib 2 fragment binds to proteins with the same Mrs. These proteins would be expected to be receptors mediating the Fib 2 fragment effects. Less
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