title: Subcellular trafficking and activity of Hyal-1 and its processed forms in murine macrophages.
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 Percoll(TM) 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.
authors: URPhyM-Laboratoire de Chimie Physiologique, NARILIS, University of Namur, Namur, Belgium.
authors: Emeline
authors: E
authors: Puissant
authors: Florentine
authors: F
authors: Gilis
authors: Sophie
authors: S
authors: Dogné
authors: Bruno
authors: B
authors: Flamion
authors: Michel
authors: M
authors: Jadot
authors: Marielle
authors: M
authors: Boonen
mesh_terms: D000818:Animals; D004705:Endocytosis / Q000502:physiology; D011992:Endosomes / Q000378:metabolism; D006031:Glycosylation; D006821:Hyaluronoglucosaminidase / Q000378:metabolism*; D006867:Hydrolases / Q000378:metabolism; D037181:Lectins, C-Type / Q000378:metabolism; D008247:Lysosomes / Q000378:metabolism; D008264:Macrophages / Q000378:metabolism*; D000090323:Mannose Receptor; D037241:Mannose-Binding Lectins / Q000378:metabolism; D008360:Mannosephosphates / Q000378:metabolism; D051379:Mice; D021381:Protein Transport / Q000502:physiology*; D011956:Receptors, Cell Surface / Q000378:metabolism; D013395:Sucrose / Q000378:metabolism
chemical_list: D037181:Lectins, C-Type; D000090323:Mannose Receptor; D037241:Mannose-Binding Lectins; D008360:Mannosephosphates; D011956:Receptors, Cell Surface; C027693:mannose-6-phosphate; D013395:Sucrose; D006867:Hydrolases; C469220:Hyal1 protein, mouse; D006821:Hyaluronoglucosaminidase
keywords: Hyal-1; lysosome; macrophages; mannose 6-phosphate; mannose receptor; maturation; secretion/recapture