Low density lipoprotein receptor-related protein mediates endocytic clearance of pro-MMP-2.TIMP-2 complex through a thrombospondin-independent mechanism.
The low density lipoprotein receptor-related protein (LRP) mediates the endocytic clearance of various proteinases and proteinase.inhibitor complexes, including thrombospondin (TSP)-dependent endocytosis of matrix metalloproteinase (MMP)-2 (or gelatinase A), a key effector of extracellular matrix remodeling and cancer progression. However, the zymogen of MMP-2 (pro-MMP-2) mostly occurs in tissues as a complex with the tissue inhibitor of MMPs (TIMP-2). Here we show that clearance of the pro-MMP-2.TIMP-2 complex is also mediated by LRP, because addition of receptor-associated protein (RAP), a natural LRP ligand antagonist, inhibited endocytosis and lysosomal degradation of (125)I-pro-MMP-2.TIMP-2. Both TIMP-2 and the pro-MMP-2 collagen-binding domain independently competed for endocytosis of (125)I-pro-MMP-2.TIMP-2 complex. Surface plasmon resonance studies indicated that pro-MMP-2, TIMP-2, and pro-MMP-2.TIMP-2 directly interact with LRP in the absence of TSP. LRP-mediated endocytic clearance of (125)I-pro-MMP-2 was inhibited by anti-TSP antibodies and accelerated upon complexing with TSP-1, but these treatments had no effect on (125)I-pro-MMP-2.TIMP-2 uptake. This implies that mechanisms of clearance by LRP of pro-MMP-2 and pro-MMP-2.TIMP-2 complex are different. Interestingly, RAP did not inhibit binding of (125)I-pro-MMP-2.TIMP-2 to the cell surface. We conclude that clearance of pro-MMP-2.TIMP-2 complex is a TSP-independent two-step process, involving (i) initial binding to the cell membrane in a RAP-insensitive manner and (ii) subsequent LRP-dependent (RAP-sensitive) internalization and degradation.
CNRS UMR 6198, IFR 53 Biomolecules, Faculty of Medicine, F-51100 Reims, France.
Hervé
H
Emonard
Georges
G
Bellon
Linda
L
Troeberg
Alix
A
Berton
Arnaud
A
Robinet
Patrick
P
Henriet
Etienne
E
Marbaix
Kirstine
K
Kirkegaard
László
L
Patthy
Yves
Y
Eeckhout
Hideaki
H
Nagase
William
W
Hornebeck
Pierre J
PJ
Courtoy
D001665:Binding Sites; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D003094:Collagen / Q000378:metabolism; D017077:Culture Media, Conditioned; D004705:Endocytosis / Q000502:physiology*; D005354:Fibrosarcoma; D015870:Gene Expression; D006801:Humans; D026503:Low Density Lipoprotein Receptor-Related Protein-1 / Q000502:physiology*; D020778:Matrix Metalloproteinase 2 / Q000235:genetics / Q000378:metabolism*; D011498:Protein Precursors / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins; D019700:Thrombospondin 1 / Q000378:metabolism / Q000494:pharmacology; D019699:Thrombospondins / Q000502:physiology*; D019716:Tissue Inhibitor of Metalloproteinase-2 / Q000235:genetics / Q000378:metabolism*; D014162:Transfection; D014407:Tumor Cells, Cultured
D017077:Culture Media, Conditioned; D026503:Low Density Lipoprotein Receptor-Related Protein-1; D011498:Protein Precursors; D011994:Recombinant Proteins; D019700:Thrombospondin 1; D019699:Thrombospondins; D019716:Tissue Inhibitor of Metalloproteinase-2; D003094:Collagen; D020778:Matrix Metalloproteinase 2