Abstract
The hyaluronidase Hyal-1 is an acid hydrolase that degrades hyaluronic acid (HA), a component of the extracellular matrix. It is often designated as a lysosomal protein. Yet few data are available on its intracellular localization and trafficking. We demonstrate here that in RAW264.7 murine macrophages, Hyal-1 is synthesized as a glycosylated precursor that is only weakly mannose 6-phosphorylated. Nevertheless, this precursor traffics to endosomes, via a mannose 6-phosphate-independent secretion/recapture mechanism that involves the mannose receptor. Once in endosomes, it is processed into a lower molecular mass form that is transported to lysosomes, where its activity could be detected using native gel zymography. Indeed, this activity co-distributed with lysosomal hydrolases in the densest fraction of a self-forming PercollTM density gradient. Moreover, it shifted toward the lower density region, in parallel with those hydrolases, when a decrease of lysosomal density was induced by the endocytosis of sucrose. Interestingly, the activity of the processed form of Hyal-1 was largely underestimated when assayed by zymography after SDS-PAGE and subsequent renaturation of the proteins, by contrast to the full-length protein that could efficiently degrade HA in those conditions. These results suggest that noncovalent associations support the lysosomal activity of Hyal-1.
Original language | English |
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Pages (from-to) | 500-15 |
Number of pages | 16 |
Journal | Traffic (Copenhagen, Denmark) |
Volume | 15 |
Issue number | 5 |
Early online date | 6 Feb 2014 |
DOIs | |
Publication status | Published - 2014 |
Keywords
- Animals
- Endocytosis
- Endosomes
- Glycosylation
- Hyaluronoglucosaminidase
- Hydrolases
- Lectins, C-Type
- Lysosomes
- Macrophages
- Mannose-Binding Lectins
- Mannosephosphates
- Mice
- Protein Transport
- Receptors, Cell Surface
- Sucrose
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Physical Chemistry and characterization(PC2)
Wouters, J. (Manager), Aprile, C. (Manager) & Fusaro, L. (Manager)
Technological Platform Physical Chemistry and characterizationFacility/equipment: Technological Platform