Abstract
Degradation of the cartilage proteoglycan aggrecan is an early event in the development of osteoarthritis, and a disintegrin and metalloproteinase with thrombospondin motifs-4 (ADAMTS-4) and ADAMTS-5 are considered to be the major aggrecan-degrading enzymes. We have recently found that ADAMTS-5 is rapidly endocytosed via low density lipoprotein receptor-related protein 1 (LRP1) and degraded by chondrocytes. Here we report that this regulatory mechanism also applies to ADAMTS-4, although its rate of endocytosis is slower than that of ADAMTS-5. Domain deletion mutagenesis of ADAMTS-4 identified that the cysteine-rich and spacer domains are responsible for binding to LRP1, whereas the thrombospondin 1 and spacer domains are responsible in ADAMTS-5. The estimated t½ value of ADAMTS-4 endocytosis was about 220 min, whereas that of ADAMTS-5 was 100 min. The difference in half-lives between the two enzymes is explained by the 13-fold lower affinity of ADAMTS-4 for LRP1 compared with that of ADAMTS-5. Studies using soluble ligand binding clusters of LRP1 showed that ADAMTS-4 binds to clusters II and IV with similar KD,app values of 98 and 73 nm, respectively, whereas ADAMTS-5 binds to cluster II, III, and IV with KD,app values of 3.5, 41, and 9 nm, respectively. Thus, ADAMTS-5 competitively inhibits ADAMTS-4 endocytosis but not vice versa. This study highlights that the affinity between a ligand and LRP1 dictates the rate of internalization and suggests that LRP1 is a major traffic controller of the two aggrecanases, especially under inflammatory conditions, where the protein levels of ADAMTS-4 increase, but those of ADAMTS-5 do not.
Original language | English |
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Pages (from-to) | 6462-6474 |
Number of pages | 13 |
Journal | The Journal of Biological Chemistry |
Volume | 289 |
Issue number | 10 |
DOIs | |
Publication status | Published - 7 Mar 2014 |
Keywords
- ADAM Proteins/chemistry
- ADAMTS4 Protein
- ADAMTS5 Protein
- Animals
- Cartilage, Articular/metabolism
- Catalytic Domain/genetics
- Cells, Cultured
- Endocytosis
- Half-Life
- Humans
- Low Density Lipoprotein Receptor-Related Protein-1/metabolism
- Osteoarthritis/metabolism
- Procollagen N-Endopeptidase/chemistry
- Protein Binding
- Sequence Deletion
- Swine