Oligomerization through Hemopexin and Cytoplasmic Domains Regulates the Activity and Turnover of Membrane-type 1 Matrix Metalloproteinase

Kaisa Lehti,Jouko Lohi,Minna M Juntunen,Duanqing Pei,Jorma Keski-Oja

doi:10.1074/jbc.m109128200

Abstract

The formation of multimeric complexes by membrane-type 1 matrix metalloproteinase (MT1-MMP) may facilitate its autocatalytic inactivation or proMMP-2 activation on the cell surface. To characterize these processes, we expressed various glutathione S-transferase/MT1-MMP fusion proteins in human HT-1080 fibrosarcoma cells and SV40-transformed lung fibroblasts and analyzed their effects on MT1-MMP activity and potential homophilic interactions. We report here that MT1-MMP is expressed on the cell surface as oligomeric 200--240-kDa complexes containing both the active 60-kDa and autocatalytically processed 43-kDa species. Overexpression of a glutathione S-transferase/MT1-MMP fusion protein containing the transmembrane and cytoplasmic domains of MT1-MMP inhibited the phorbol 12-myristate 13-acetate-induced autocatalytic cleavage of endogenous MT1-MMP to the 43-kDa species, but not proMMP-2 activation. On the other hand, a similar fusion protein with the hemopexin, transmembrane, and cytoplasmic domains inhibited proMMP-2 activation in a dominant-negative fashion. These results suggest that both the autocatalytic cleavage of MT1-MMP and proMMP-2 activation may be regulated by oligomerization through the cytoplasmic and hemopexin domains. Indeed, either domain, when attached to the cell membrane by a transmembrane domain, formed stable homophilic complexes. Copurification of MT1-MMP with these fusion proteins correlated with their cell-surface co-localization. Thus, MT1-MMP oligomerization through the hemopexin, transmembrane, and cytoplasmic domains controls its catalytic activity.

Highlights

The formation of multimeric complexes by membranetype 1 matrix metalloproteinase (MT1-MMP) may facilitate its autocatalytic inactivation or proMMP-2 activation on the cell surface
Dissociation of the cross-linked complexes by reduction revealed that the cell-surface complexes of PMAtreated or MT1-MMP-overexpressing HT-1080 cells were composed of both the active 60-kDa and proteolytically inactive 43-kDa species (Fig. 2A, lower panel, lanes 2– 4), arguing that these MT1-MMP species are present on the cell surface as high molecular mass complexes
Our results indicate that overexpression of non-catalytic mutants capable of interacting with wild-type MT1-MMP through the cytoplasmic, transmembrane, and extracellular hemopexin domains leads to competitive inhibition of MMP-2 activation by wild-type MT1-MMP molecules in a dominantnegative fashion

Summary

Introduction

The formation of multimeric complexes by membranetype 1 matrix metalloproteinase (MT1-MMP) may facilitate its autocatalytic inactivation or proMMP-2 activation on the cell surface. On the HT-1080 cell surface, cross-linked endogenous MT1-MMP was detected under nonreducing conditions as 200 –240-kDa complexes (Fig. 2A, upper panel, lane 1).

Results

Conclusion