Siglec-15 Regulates Osteoclast Differentiation by Modulating RANKL-Induced Phosphatidylinositol 3-Kinase/Akt and Erk Pathways in Association With Signaling Adaptor DAP12

Abstract

Siglecs are a family of sialic acid-binding immunoglobulin-like lectins that regulate the functions of cells in the innate and adaptive immune systems through glycan recognition. Here we show that Siglec-15 regulates osteoclast development and bone resorption by modulating receptor activator of nuclear factor kB ligand (RANKL) signaling in association with DNAX-activating protein 12 kDa (DAP12), an adaptor protein bearing an immunoreceptor tyrosine-based activation motif (ITAM). Among the known Siglecs expressed in myeloid lineage cells, only Siglec-15 was upregulated by RANKL in mouse primary bone marrow macrophages. Siglec-15-deficient mice exhibit mild osteopetrosis resulting from impaired osteoclast development. Consistently, cells lacking Siglec-15 exhibit defective osteoclast development and resorptive activity in vitro. RANKL-induced activation of phosphatidylinositol 3-kinase (PI3K)/Akt and Erk pathways were impaired in Siglec-15-deficient cells. Retroviral transduction of Siglec-15-null osteoclast precursors with wild-type Siglec-15 or mutant Siglec-15 revealed that the association of Siglec-15 with DAP12 is involved in the downstream signal transduction of RANK. Furthermore, we found that the ability of osteoclast formation is preserved in the region adjacent to the growth plate in Siglec-15-deficient mice, indicating that there is a compensatory mechanism for Siglec-15-mediated osteoclastogenesis in the primary spongiosa. To clarify the mechanism of this compensation, we examined whether osteoclast-associated receptor (OSCAR)/Fc receptor common g (FcRg) signaling, an alternative ITAM-mediated signaling pathway to DAP12, rescues impaired osteoclastogenesis in Siglec-15-deficient cells. The ligands in type II collagen activate OSCAR and rescue impaired osteoclastogenesis in Siglec-15-deficient cells when cultured on bone slices, indicating that Siglec-15-mediated signaling can be compensated for by signaling activated by type II collagen and other bone matrix components in the primary spongiosa. Our findings indicate that Siglec-15 plays an important role in physiologic bone remodeling by modulating RANKL signaling, especially in the secondary spongiosa.

Mechanism and Function of Monoclonal Antibodies Targeting Siglec-15 for Therapeutic Inhibition of Osteoclastic Bone Resorption

“…This could potentially result in a reduction in the rare but devastating side effects of current therapeutics, which include osteonecrosis of the jaw (39), and atypical fractures of the femur (40). Both the specificity of Siglec-15 expression in osteoclasts and the potential for Siglec-15 targeted therapies to preserve osteoblast-osteoclast coupling and bone formation are supported by the mildly osteopetrotic but otherwise normal phenotype of the recently described siglec-15 Ϫ/Ϫ mice (27,28). Although further studies will be helpful to fully characterize their in vivo effects, these characteristics suggest that Siglec-15 antibodies could have distinct advantages over existing drugs, and therefore, we believe that continued development of this novel class of bone loss therapeutic is certainly warranted.…”

“…Very recently, following completion of the current study, two groups have independently generated full-body siglec-15 Ϫ/Ϫ mice (27,28). These mice are viable and healthy and display a mildly osteopetrotic phenotype.…”

Sialic acid-binding immunoglobulin-like lectin 15 (Siglec-15) mediates periarticular bone loss, but not joint destruction, in murine antigen-induced arthritis

“…Sialic acid-binding Ig-like lectin (Siglec)-15 is a DAP12-associated immunoreceptor that recognizes sialylated glycans and regulates osteoclast differentiation [11,12]. We previously demonstrated that Siglec-15 is involved in physiologic bone remodeling by modulating RANKL signaling [13].…”

Glucosamine Suppresses Osteoclast Differentiation through the Modulation of Glycosylation Including <i>O</i>-GlcNAcylation

“…13) In addition, sugar-binding proteins such as siglec, which interact with sialic acid, and galectins, which interact with Galβ1-4GlcNAc, also have important roles in osteoclastogenesis. [14][15][16][17][18][19] Therefore, in order to clarify the potential for GlcN-mediated glycosylation modification, we investigated the effect of GlcN treatment on the pattern of glycosylation using lectin blotting of cell extracts with SSA, MAM, RCA120, PHA-E 4 , Con A, and LCA, which mainly recognize α2,6-linked sialic acid, α2,3-linked sialic acid, Galβ1-4GlcNAc, bisecting GlcNAc, high mannose type Nglycan, and N-glycan core modified with fucose, respectively (Figs. 2B-G).…”