Protein kinase N3 promotes bone resorption by osteoclasts in response to Wnt5a-Ror2 signaling

Abstract

Cytoskeletal reorganization in osteoclasts to form actin rings is necessary for these cells to attach to bone and resorb bone matrices. We delineated the pathway through which Wnt5a signaling through receptor tyrosine kinase-like orphan receptor 2 (Ror2) promoted the bone-resorbing activity of osteoclasts. Wnt5a binding to Ror2 stimulated Rho, a small GTPase involved in cytoskeletal reorganization. Subsequently, the Rho effector kinase Pkn3 bound to and enhanced the activity of c-Src, a nonreceptor tyrosine kinase that is critical for actin ring formation. Mice with an osteoclast-specific deficiency in () had increased bone mass. Osteoclasts derived from these mice exhibited impaired bone resorption and actin ring formation, defects that were rescued by overexpression of constitutively active RhoA. These osteoclasts also exhibited reduced interaction between c-Src and Pkn3 and reduced c-Src kinase activity. Similar to mice, mice with a global deficiency of () had increased bone mass. The proline-rich region and kinase domain of Pkn3 were required to restore the bone-resorbing activity of osteoclasts derived from mice. Thus, Pkn3 promotes bone resorption downstream of Wnt5a-Ror2-Rho signaling, and this pathway may be a therapeutic target for bone diseases such as osteoporosis, rheumatoid arthritis, and periodontal disease.

Deletion of Wnt5a in osteoclasts results in bone loss through decreased bone formation

“…WNT5A is the prototypical noncanonical WNT, which when produced by osteoblasts controls the BMU by stimulating the expression of RANK in osteoclast precursors . Osteoclast precursors also express a basal level of WNT5A that mirrors the biological effects of WNT5A from osteoblast‐lineage cells where it promotes osteoclastogenesis through an autocrine mechanism . Our in vitro experiments are in agreement with these earlier findings, that in the absence of osteoblasts there were fewer osteoclasts and decreased osteoclast activity when Wnt5a was deleted from osteoclast precursors.…”

“…While WNT5A is known to undergo extensive lipidation and glycosylation before secretion, reports of phosphorylation have not been described previously. This finding is also intriguing as it corresponds to the increase in Wnt5a gene and protein expression that occurs during osteoclastogenesis . This uniquely phosphorylated WNT5A from osteoclast‐like cells may prove to have an important role in controlling osteoclast and osteoblast dynamics during bone remodeling.…”

“…Expression of Wnt5a in osteoclast precursors also increases in a time‐dependent manner in response to RANKL‐stimulation, which suggests that osteoclast‐derived WNT5A may act as a clastokine to couple osteoclast activity to that of osteoblasts within the BMU. When we targeted Wnt5a in mature osteoclasts in vivo using Ctsk ‐ Cre , we observed a low bone–mass phenotype that was present in multiple sites, including the femur and vertebral body.…”

“…Ctsk ‐ Cre mice were bred with the Wnt5a F/F mice to generate Ctsk ‐ Cre ; Wnt5a F/+ mice, which were then crossed with Wnt5a F/F mice to generate experimental Ctsk ‐ Cre ; Wnt5a F/F ( Wnt5a cKO) mice. Ctsk‐Cre is a knock‐in mutation that is under the control of the endogenous promoter of the Ctsk gene and has previously been shown to effectively target floxed genes in mature osteoclasts . Conditional knockout mice were compared with Wnt5a F/+ littermate controls.…”

“…Genetic studies manipulating the orphan receptor Ror2 in mice established that noncanonical WNT signaling in osteoclast precursors increases bone mass through a mechanism linked to decreased osteoclastogenesis . WNT5A is a noncanonical WNT secreted by osteoblasts and, to a lesser extent, osteoclasts that mediates cross‐communication between the two distinct cell types.…”