Osteoclasts are the principal cells that efficiently resorb bone. Numerous studies have attempted to reveal the molecular pathways leading to the differentiation and activation of osteoclasts to ...improve the treatment and prevention of osteoporosis and other bone-destructive diseases. While the cumulative knowledge of osteoclast regulatory molecules, such as receptor activator of nuclear factor-kB ligand (RANKL) and nuclear factor of activated T cells 1 (NFATc1), contributes to the understanding of the developmental progression of osteoclasts, little is known about how the discrete steps of osteoclastogenesis modify osteoclast status but not the absolute number of osteoclasts. The regulatory mechanisms involved in osteoclast maturation but not those involved in differentiation deserve special attention due to their potential use in establishing a more effective treatment strategy: targeting late-phase differentiation while preserving coupled bone formation. Recent studies have shed light on the molecules that govern late-phase osteoclast differentiation and maturation, as well as the metabolic changes needed to adapt to shifting metabolic demands. This review outlines the current understanding of the regulation of osteoclast differentiation, as well as osteoclast metabolic adaptation as a differentiation control mechanism. Additionally, this review introduces molecules that regulate the late-phase osteoclast differentiation and thus minimally impact coupled bone formation.
Protocadherin-7 (Pcdh7) is a member of the non-clustered protocadherin δ1 subgroup of the cadherin superfamily. Although the cell-intrinsic role of Pcdh7 in osteoclast differentiation has been ...demonstrated, the molecular mechanisms of Pcdh7 regulating osteoclast differentiation remain to be determined. Here, we demonstrate that Pcdh7 contributes to osteoclast differentiation by regulating small GTPases, RhoA and Rac1, through its SET oncoprotein binding domain. Pcdh7 is associated with SET along with RhoA and Rac1 during osteoclast differentiation. Pcdh7-deficient (Pcdh7
) cells showed abolished RANKL-induced RhoA and Rac1 activation, and impaired osteoclast differentiation. Impaired osteoclast differentiation in Pcdh7
cells was restored by retroviral transduction of full-length Pcdh7 but not by a Pcdh7 mutant that lacks SET binding domain. The direct crosslink of the Pcdh7 intracellular region induced the activation of RhoA and Rac1, which was not observed when Pcdh7 lacks the SET binding domain. Additionally, retroviral transduction of the constitutively active form of RhoA and Rac1 completely restored the impaired osteoclast differentiation in Pcdh7
cells. Collectively, these results demonstrate that Pcdh7 controls osteoclast differentiation by regulating RhoA and Rac1 activation through the SET binding domain.
Osteoclasts are primary bone-resorbing cells, and receptor-activated NF-kB ligand (RANKL) stimulation is the key driver of osteoclast differentiation. During late-stage differentiation, osteoclasts ...become multinucleated and enlarged (so-called "maturation"), suggesting their need to adapt to changing metabolic demands and a substantial increase in size. Here, we demonstrate that immunoglobulin superfamily 11 (IgSF11), which is required for osteoclast differentiation through an association with the postsynaptic scaffolding protein PSD-95, regulates osteoclast differentiation by controlling the activity of pyruvate kinase M isoform 2 (PKM2). By using a system that directly induces the activation of IgSF11 in a controlled manner, we identified PKM2 as a major IgSF11-induced tyrosine-phosphorylated protein. IgSF11 activates multiple Src family tyrosine kinases (SFKs), including c-Src, Fyn, and HcK, which phosphorylate PKM2 and thereby inhibit PKM2 activity. Consistently, IgSF11-deficient cells show higher PKM2 activity and defective osteoclast differentiation. Furthermore, inhibiting PKM2 activities with the specific inhibitor Shikonin rescues the impaired osteoclast differentiation in IgSF11-deficient cells, and activating PKM2 with the specific activator TEPP46 suppresses osteoclast differentiation in wild-type cells. Moreover, PKM2 activation further suppresses osteoclastic bone loss without affecting bone formation in vivo. Taken together, these results show that IgSF11 controls osteoclast differentiation through PKM2 activity, which is a metabolic switch necessary for optimal osteoclast maturation.
Protocadherin-7 (Pcdh7) is a member of the non-clustered protocadherin δ1 subgroup of the cadherin superfamily. Pcdh7 has been revealed to control osteoclast differentiation by regulating Rho-family ...small GTPases, RhoA and Rac1, through its intracellular SET binding domain. However, the mechanisms by which small GTPases are regulated downstream of Pcdh7 remain unclear. Here, we demonstrate that protein phosphatase 2A (PP2A)-mediated dephosphorylation of Glycogen synthase kinase-3β (GSK3β) is required for Pcdh7-dependent activation of RhoA during osteoclast differentiation. Pcdh7-deficient (Pcdh7
) cells showed impaired PP2A activity, despite their normal expression of PP2A. GSK3β, whose activity is regulated by its inhibitory phosphorylation at Ser9, was dephosphorylated during osteoclast differentiation in a Pcdh7-dependent manner. Inhibition of protein phosphatase by okadaic acid reduced dephosphorylation of GSK3β in Pcdh7
cells, while activation of PP2A by DT-061 rescued impaired dephosphorylation of GSK3β in Pcdh7
cells. Inhibition of GSK3β by AR-A014418 inhibited RANKL-induced RhoA activation and osteoclast differentiation in Pcdh7
cells. On the other hand, DT-061 treatment rescued impaired RhoA activation and RANKL-induced osteoclast differentiation in Pcdh7
cells. Taken together, these results demonstrate that PP2A dephosphorylates GSK3β and thereby activates it in a Pcdh7-dependent manner, which is required for activation of small GTPase RhoA and proper osteoclast differentiation.
At the optic chiasm, retinal ganglion cells (RGCs) project ipsi- or contralaterally to establish the circuitry for binocular vision. Ipsilateral guidance programs have been characterized, but ...contralateral guidance programs are not well understood. Here, we identify a tripartite molecular system for contralateral RGC projections: Semaphorin6D (Sema6D) and Nr-CAM are expressed on midline radial glia and Plexin-A1 on chiasm neurons, and Plexin-A1 and Nr-CAM are also expressed on contralateral RGCs. Sema6D is repulsive to contralateral RGCs, but Sema6D in combination with Nr-CAM and Plexin-A1 converts repulsion to growth promotion. Nr-CAM functions as a receptor for Sema6D. Sema6D, Plexin-A1, and Nr-CAM are all required for efficient RGC decussation at the optic chiasm. These findings suggest a mechanism by which a complex of Sema6D, Nr-CAM, and Plexin-A1 at the chiasm midline alters the sign of Sema6D and signals Nr-CAM/Plexin-A1 receptors on RGCs to implement the contralateral RGC projection.
► Plexin-A1 and Nr-CAM are expressed in contralateral RGCs and the optic chiasm midline ► Nr-CAM and Plexin-A1 modulate inhibition by midline Sema6D ► Nr-CAM is a receptor for Sema6D and binds to Plexin-A1 ► Proper RGC axon fasciculation and decussation are dependent on all three factors
Kuwajima et al. identify molecular interactions mediating the contralateral retinal ganglion cell projection. A complex of Nr-CAM and Plexin-A1 and Semaphorin6D at the optic chiasm switches the responsiveness of RGCs to inhibitory Sema6D, enabling them to cross the midline.
Differentiation of osteoclasts, which are specialized multinucleated macrophages capable of bone resorption, is driven primarily by receptor activator of NF-κB ligand (RANKL). Additional signaling ...from cell surface receptors, such as cell adhesion molecules (CAMs), is also required for osteoclast maturation. Previously, we have demonstrated that immunoglobulin superfamily 11 (IgSF11), a member of the immunoglobulin-CAM (IgCAM) family, plays an important role in osteoclast differentiation through association with the scaffold protein postsynaptic density protein 95 (PSD-95). Here, we demonstrate that the osteoclast-expressed CAM CD44 can compensate for IgSF11 deficiency when cell-cell interaction conditions are suboptimal by associating with PSD-95. Impaired osteoclast differentiation in IgSF11-deficient (IgSF11
) cultures was rescued by antibody-mediated stimulation of CD44 or by treatment with low-molecular-weight hyaluronan (LMW-HA), a CD44 ligand. Biochemical analysis revealed that PSD-95, which is required for osteoclast differentiation, associates with CD44 in osteoclasts regardless of the presence or absence of IgSF11. RNAi-mediated knockdown of PSD-95 abrogated the effects of either CD44 stimulation or LMW-HA treatment on osteoclast differentiation, suggesting that CD44, similar to IgSF11, is functionally associated with PSD-95 during osteoclast differentiation. Taken together, these results reveal that CD44 can compensate for IgSF11 deficiency in osteoclasts through association with PSD-95.
Excessive bone resorption by osteoclasts (OCs) can result in serious clinical outcomes, including bone loss that may weaken skeletal or periodontal strength. Proper bone homeostasis and skeletal ...strength are maintained by balancing OC function with the bone-forming function of osteoblasts. Unfortunately, current treatments that broadly inhibit OC differentiation or function may also interfere with coupled bone formation. We therefore identified a factor, the purinergic receptor P2X5 that is highly expressed during the OC maturation phase, and which we show here plays no apparent role in early bone development and homeostasis, but which is required for osteoclast-mediated inflammatory bone loss and hyper-multinucleation of OCs. We further demonstrate that P2X5 is required for ATP-mediated inflammasome activation and IL-1β production by OCs, and that P2X5-deficient OC maturation is rescued in vitro by addition of exogenous IL-1β. These findings identify a mechanism by which OCs react to inflammatory stimuli, and may identify purinergic signaling as a therapeutic target for bone loss-related inflammatory conditions.
Purinergic receptor signaling is increasingly recognized as an important regulator of inflammation. The P2X family purinergic receptors P2X5 and P2X7 have both been implicated in bone biology, and it ...has been suggested recently that P2X5 may be a significant regulator of inflammatory bone loss. However, a role for P2X5 in periodontitis is unknown. The present study aimed to evaluate the functional role of P2X5 in ligatureinduced periodontitis in mice. Five days after placement of ligature, analysis of alveolar bone revealed decreased bone loss in P2rx5-/- mice compared to P2rx7-/- and WT control mice. Gene expression analysis of the gingival tissue of ligated mice showed that IL1b, IL6, IL17a and Tnfsf11 expression levels were significantly reduced in P2rx5-/- compared to WT mice. These results suggest the P2X5 receptor may regulate bone loss related to periodontitis and it may thus be a novel therapeutic target in this oral disease. BMB Reports 2018; 51(9): 468-473.
Osteoclasts are hematopoietic-derived cells that resorb bone. They are required to maintain proper bone homeostasis and skeletal strength. Although osteoclast differentiation depends on receptor ...activator of NF-κB ligand (RANKL) stimulation, additional molecules further contribute to osteoclast maturation. Here, we demonstrate that protocadherin-7 (Pcdh7) regulates formation of multinucleated osteoclasts and contributes to maintenance of bone homeostasis. We found that Pcdh7 expression is induced by RANKL stimulation, and that RNAi-mediated knockdown of Pcdh7 resulted in impaired formation of osteoclasts. We generated Pcdh7-deficient mice and found increased bone mass due to decreased bone resorption but without any defect in bone formation. Using an in vitro culture system, it was revealed that formation of multinucleated osteoclasts is impaired in Pcdh7-deficient cultures, while no apparent defects were observed in differentiation and function of Pcdh7-deficient osteoblasts. Taken together, these results reveal an osteoclast cell-intrinsic role for Pcdh7 in maintaining bone homeostasis. BMB Reports 2020; 53(9): 472-477.
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