Abstract
Osteoclasts are giant bone-resorbing cells originating from monocytes/macrophages. During their differentiation, they overexpress two lysosomal enzymes, cathepsin K and TRAP, which are secreted into the resorption lacuna, an acidified sealed area in contact with bone matrix where bone degradation takes place. Here we report that the acid hydrolase HYAL1, a hyaluronidase able to degrade the glycosaminoglycans hyaluronic acid (HA) and chondroitin sulfate, is also upregulated upon osteoclastogenesis. The mRNA expression and protein level of HYAL1 are markedly increased in osteoclasts differentiated from RAW264.7 mouse macrophages or primary mouse bone marrow monocytes compared to these precursor cells. As a result, the HYAL1-mediated HA hydrolysis ability of osteoclasts is strongly enhanced. Using subcellular fractionation, we demonstrate that HYAL1 proteins are sorted to the osteoclast lysosomes even though, in contrast to cathepsin K and TRAP, HYAL1 is poorly mannose 6-phosphorylated. We reported previously that macrophages secrete HYAL1 proforms by constitutive secretion, and that these are recaptured by the cell surface mannose receptor, processed in endosomes and sorted to lysosomes. Present work highlights that osteoclasts secrete HYAL1 in two ways, through lysosomal exocytosis and constitutive secretion, and that these cells promote the extracellular residency of HYAL1 through downregulation of the mannose receptor. Interestingly, the expression of the other main hyaluronidase, HYAL2, and of lysosomal exoglycosidases involved in HA degradation, does not increase similarly to HYAL1 upon osteoclastogenesis. Taken together, these findings point out the predominant involvement of HYAL1 in bone HA metabolism and perhaps bone remodeling via the resorption lacuna.
Highlights
Osteoclasts are giant multinucleated cells responsible for bone resorption that differentiate from hematopoietic cells of monocyte/macrophage lineage
As expected (Fig 1A), we observed that differentiated osteoclasts contain large amounts of mature/proteolytically processed forms of cathepsin K and TRAP (Fig 1A, open arrowheads), as well as low amounts of their precursor forms, whereas macrophages and cells treated for 2 days with receptor activator of nuclear factor kappa-B ligand (RANKL) are mostly devoid of these proteins
We previously reported that RAW264.7 macrophages contain two main forms of HYAL1 that are detected by western blotting under reducing conditions: a ~52 kDa precursor form that bears high-mannose N-linked glycans and likely localizes to the endoplasmic reticulum (ER), and a mature form of ~48 kDa produced by proteolytic cleavage [26]
Summary
Osteoclasts are giant multinucleated cells responsible for bone resorption that differentiate from hematopoietic cells of monocyte/macrophage lineage. When in contact with bone, osteoclasts polarize and form a resorption lacuna sealed by an actin ring in which the bone matrix is degraded, notably by secreted lysosomal acid hydrolases [1,2]. These are released when secretory lysosomes fuse with the osteoclast apical membrane, a process that provides vATPase complexes which generate an acidified extracellular environment optimal for lysosomal hydrolase activity [3,4,5,6,7]. One common feature of cathepsin K and TRAP is their marked upregulation, in contrast to other lysosomal acid hydrolases, upon differentiation of osteoclasts from their monocytic precursors [17]
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