title: Siglec-15 protein regulates formation of functional osteoclasts in concert with DNAX-activating protein of 12 kDa (DAP12).
abstract: Osteoclasts are multinucleated giant cells that reside in osseous tissues and resorb bone. Signaling mediated by receptor activator of nuclear factor (NF)-κB (RANK) and its ligand leads to the nuclear factor of activated T cells 2/c1 (NFAT2 or NFATc1) expression, a critical step in the formation of functional osteoclasts. In addition, adaptor proteins harboring immunoreceptor tyrosine-based activation motifs, such as DNAX-activating protein of 12 kDa (DAP12), play essential roles. In this study, we identified the gene encoding the lectin Siglec-15 as NFAT2-inducible, and we found that the protein product links RANK ligand-RANK-NFAT2 and DAP12 signaling in mouse osteoclasts. Both the recognition of sialylated glycans by the Siglec-15 V-set domain and the association with DAP12 through its Lys-272 are essential for its function. When Siglec-15 expression was knocked down, fewer multinucleated cells developed, and those that did were morphologically contracted with disordered actin-ring structures. These changes were accompanied by significantly reduced bone resorption. Siglec-15 formed complexes with Syk through DAP12 in response to vitronectin. Furthermore, chimeric molecules consisting of the extracellular and transmembrane regions of Siglec-15 with a K272A mutation and the cytoplasmic region of DAP12 significantly restored bone resorption in cells with knocked down Siglec-15 expression. Together, these results suggested that the Siglec-15-DAP12-Syk-signaling cascade plays a critical role in functional osteoclast formation.
authors.affiliation: Graduate School of Biological Sciences, Nara Institute of Science and Technology, Ikoma, Nara 630-0192. Electronic address: n-ishida@bs.naist.jp.
authors.forename: Norihiro
authors.initials: N
authors.lastname: Ishida-Kitagawa
authors.affiliation: Graduate School of Biological Sciences, Nara Institute of Science and Technology, Ikoma, Nara 630-0192.
authors.forename: Kunitaro
authors.initials: K
authors.lastname: Tanaka
authors.affiliation: Graduate School of Biological Sciences, Nara Institute of Science and Technology, Ikoma, Nara 630-0192.
authors.forename: Xilinqiqige
authors.initials: X
authors.lastname: Bao
authors.affiliation: Graduate School of Biological Sciences, Nara Institute of Science and Technology, Ikoma, Nara 630-0192.
authors.forename: Takanori
authors.initials: T
authors.lastname: Kimura
authors.affiliation: Graduate School of Biological Sciences, Nara Institute of Science and Technology, Ikoma, Nara 630-0192.
authors.forename: Tadashi
authors.initials: T
authors.lastname: Miura
authors.affiliation: Graduate School of Biological Sciences, Nara Institute of Science and Technology, Ikoma, Nara 630-0192.
authors.forename: Yoshiki
authors.initials: Y
authors.lastname: Kitaoka
authors.affiliation: Graduate School of Biological Sciences, Nara Institute of Science and Technology, Ikoma, Nara 630-0192.
authors.forename: Kouhei
authors.initials: K
authors.lastname: Hayashi
authors.affiliation: Graduate School of Biological Sciences, Nara Institute of Science and Technology, Ikoma, Nara 630-0192; Research Institute for Microbial Diseases, Osaka University, Osaka 565-0871.
authors.forename: Mizuho
authors.initials: M
authors.lastname: Sato
authors.affiliation: Graduate School of Biological Sciences, Nara Institute of Science and Technology, Ikoma, Nara 630-0192; Laboratory of Single-Molecule Cell Biology, Tohoku University Graduate School of Life Sciences, Aoba-ku, Sendai, Miyagi 980-8578.
authors.forename: Masahiro
authors.initials: M
authors.lastname: Maruoka
authors.affiliation: Graduate School of Biological Sciences, Nara Institute of Science and Technology, Ikoma, Nara 630-0192.
authors.forename: Takuya
authors.initials: T
authors.lastname: Ogawa
authors.affiliation: Department of Molecular Biology, Osaka Medical Center for Cancer and Cardiovascular Diseases, Higashinari-ku, Osaka, 537-8511, Japan.
authors.forename: Jun
authors.initials: J
authors.lastname: Miyoshi
authors.affiliation: Graduate School of Biological Sciences, Nara Institute of Science and Technology, Ikoma, Nara 630-0192.
authors.forename: Tatsuo
authors.initials: T
authors.lastname: Takeya
mesh_terms: D048868:Adaptor Proteins, Signal Transducing / Q000096:biosynthesis* / Q000235:genetics; D000818:Animals; D001862:Bone Resorption / Q000235:genetics / Q000378:metabolism; D016466:CHO Cells; D006224:Cricetinae; D003412:Cricetulus; D005786:Gene Expression Regulation / Q000502:physiology; D008562:Membrane Glycoproteins / Q000096:biosynthesis* / Q000235:genetics; D051379:Mice; D020125:Mutation, Missense; D050778:NFATC Transcription Factors / Q000235:genetics / Q000378:metabolism; D041681:NIH 3T3 Cells; D010010:Osteoclasts / Q000166:cytology / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D053245:RANK Ligand / Q000235:genetics / Q000378:metabolism; D053246:Receptor Activator of Nuclear Factor-kappa B / Q000235:genetics / Q000378:metabolism; D011971:Receptors, Immunologic / Q000096:biosynthesis* / Q000235:genetics; D063267:Sialic Acid Binding Ig-like Lectin 1; D015398:Signal Transduction / Q000502:physiology*
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D008562:Membrane Glycoproteins; D050778:NFATC Transcription Factors; C494005:Nfatc1 protein, mouse; D053245:RANK Ligand; D053246:Receptor Activator of Nuclear Factor-kappa B; D011971:Receptors, Immunologic; D063267:Sialic Acid Binding Ig-like Lectin 1; C571167:Siglec1 protein, mouse; C505687:Tnfrsf11a protein, mouse; C505689:Tnfsf11 protein, mouse; C484963:Tyrobp protein, mouse