Abstract
Proprotein convertases play an important role in tumorigenesis and invasiveness. Here, we report that a dibasic amino acid convertase, furin, directly cleaves proMMP-2 within the trans-Golgi network leading to an inactive form of matrix metalloproteinase-2 (MMP-2). Co-transfection of COS-1 cells with both proMMP-2 and furin cDNAs resulted in the cleavage of the N-terminal propeptide of proMMP-2. The molecular mass of cleaved MMP-2 (63 kDa), detected in both cell lysates and conditioned medium, is between the intermediate and fully activated forms of MMP-2 induced by membrane type 1-MMP. Furin-cleaved MMP-2 does not possess proteolytic activity as examined in a cell-free assay. Treatment of transfected cells with a furin inhibitor resulted in a dose-dependent inhibition of proMMP-2 cleavage; recombinant tissue inhibitor of metalloproteinase-2, which binds to the active site of membrane type 1-MMP, had no inhibitory effect. Site-directed mutagenesis of amino acids in the furin consensus recognition motif of proMMP-2(R69KPR72) prevented propeptide cleavage, thereby identifying the scissile bond and characterizing the basic amino acids required for cleavage. Other experimental observations were consistent with intracellular furin cleavage of proMMP-2 in the trans-Golgi network. The furin cleavage site in other proMMPs was examined. MMP-3, which contains the RXXR furin consensus sequence, was cleaved in furin co-transfected cells, whereas MMP-1, which lacks an RXXR consensus sequence, was not cleaved. In conclusion, we report the novel observation that furin can directly cleave the RXXR amino acid sequence in the propeptide domain of proMMP-2 leading to inactivation of the enzyme.
Highlights
Matrix metalloproteinases (MMPs)1 are an important family of zinc- and calcium-dependent proteinases that degrade extracellular matrix components and numerous other proteins
To examine the role of furin in the furin-MT1-MMP-MMP-2 activation axis, COS-1 cells were co-transfected with both furin and proMMP-2 cDNAs, and the conditioned medium from the transfected cells was examined by gelatin zymography
We present biochemical evidence that furin is capable of cleaving proMMP-2 in the trans-Golgi network (TGN) before secretion resulting in an inactive form of MMP-2
Summary
Matrix metalloproteinases (MMPs) are an important family of zinc- and calcium-dependent proteinases that degrade extracellular matrix components and numerous other proteins. The zymogens of most MMPs are activated through a two-step activation mechanism Activator proteinases, such as trypsin or plasmin or organomercurial chemical treatment, first attack the proteinase-susceptible “bait” region located in the middle of the propeptide domain [4, 5]. This cleavage induces conformational changes in the propeptide and renders the final activation site to be readily cleaved by a second proteolysis. In contrast to other secreted MMPs, proMMP-2 is physiologically activated on the cell surface through a MT-MMP-dependent mechanism [7]. Furin is a type I membrane protein localized to the TGN, a late Golgi structure that is responsible
Sign-in/Register to view highlights & summary Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
