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Martin Roderfeld, Jrgen Graf, Bernd Giese, Rebeca Salguero-Palacios, Annette Tschuschner, Gerhard Mller-Newen, Elke Roeb
Latent MMP-9 is bound to TIMP-1 before secretion
Keywords: confocal laser scanning microscopy, fluorescence resonance energy transfer, Matrix metalloproteinase, tissue inhibitor of metalloproteinases
Expression patterns of matrix metalloproteinase-9 (MMP-9) and its specific inhibitor, tissue inhibitor of metalloproteinases-1 (TIMP-1), are closely correlated with physiological and pathological processes characterized by degradation and accumulation of extracellular matrix (ECM). Both, activated MMP-9 and pro-MMP- 9 can bind to TIMP-1 and most cell types secrete MMP-9 in complex with TIMP-1. Utilizing immunofluorescence we observed intracellular co-localization of MMP-9 and TIMP-1 in stimulated human fibrosarcoma cells.
In the present study we searched for the origin of the complex formation between the latent enzyme and its specific inhibitor on subcellular level. Fluorescence resonance energy transfer (FRET) between the fluorescently labelled enzyme and its inhibitor in co-transfected cells were measured. MMP-9 and TIMP-1 were fused to cyan and yellow variants of the green fluorescent protein and transiently expressed in human hepatoma cells. The intracellular distribution of fluorescently labelled TIMP-1 and MMP-9 was analyzed by confocal laser scanning microscopy. Intracellular complex formation in the Golgi apparatus was verified demonstrating FRET between MMP-9- CFP and TIMP-1-YFP. Our data provide evidence that the proMMP-9-TIMP-1 complex is already present in the Golgi apparatus. This may be of significance for a number of intracellular and extracellular biochemical processes involving proMMP-9. However, the magnitude and functional relevance of this finding remains yet unknown.
Biological Chemistry, Walter de Gruyter
Print ISSN: 1431-6730
Volume: 2007
Pages: -