Receptor activator of NF-kappaB (RANK) ligand induces ectodomain shedding of RANK in murine RAW264.7 macrophages.
Osteoclastogenesis is a highly sophisticated process that involves a variety of membrane-bound proteins expressed in osteoblasts and osteoclast precursors. Over the past several years, proteolytic cleavage and release of the ectodomain of membrane-bound proteins, also referred to as ectodomain shedding, has emerged as an important posttranslational regulatory mechanism for modifying the function of cell surface proteins. In line with this notion, several membrane-bound molecules involved in osteoclastogenesis, including CSF-1R and receptor activator of NF-kappaB ligand (RANKL), are proteolytically cleaved and released from the cell surface. In this study, we investigated whether receptor activator of NF-kappaB (RANK), one of the most essential molecules in osteoclastogenesis, undergoes ectodomain shedding. The results showed that RANK is released in the form of a soluble monomeric protein and that TNF-alpha-converting enzyme is involved in this activity. We also identified potential cleavage sites in the juxtamembrane domain of RANK and found that rRANKL induces RANK shedding in a macrophage-like cell line RAW264.7 via TNFR-associated factor 6 and MAPK pathways. Furthermore, we found that RANKL-induced osteoclastogenesis is accelerated in TNF-alpha-converting enzyme-deficient osteoclast precursors. These observations suggest the potential involvement of ectodomain shedding in the regulation of RANK functions and may provide novel insights into the mechanisms of osteoclastogenesis.
Hakozaki
Akihiro
A
Department of Orthopedic Surgery, School of Medicine, Keio University, Tokyo 160-8582, Japan.
Yoda
Masaki
M
Tohmonda
Takahide
T
Furukawa
Mitsuru
M
Hikata
Tomohiro
T
Uchikawa
Shinichi
S
Takaishi
Hironari
H
Matsumoto
Morio
M
Chiba
Kazuhiro
K
Horiuchi
Keisuke
K
Toyama
Yoshiaki
Y
D051722:ADAM Proteins / Q000172:deficiency / Q000235:genetics / Q000378:metabolism*; D000072198:ADAM17 Protein; D000818:Animals; D001665:Binding Sites; D015153:Blotting, Western; D019556:COS Cells; D002460:Cell Line; D002522:Chlorocebus aethiops; D005434:Flow Cytometry; D008264:Macrophages / Q000166:cytology / Q000378:metabolism*; D008565:Membrane Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D051379:Mice; D018345:Mice, Knockout; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D050778:NFATC Transcription Factors / Q000235:genetics / Q000378:metabolism; D010010:Osteoclasts / Q000166:cytology / Q000378:metabolism; D053245:RANK Ligand / Q000235:genetics / Q000378:metabolism*; D053246:Receptor Activator of Nuclear Factor-kappa B / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012995:Solubility; D048029:TNF Receptor-Associated Factor 6 / Q000235:genetics / Q000378:metabolism; D014162:Transfection; D015854:Up-Regulation
D008565:Membrane Proteins; D050778:NFATC Transcription Factors; C494005:Nfatc1 protein, mouse; D053245:RANK Ligand; D053246:Receptor Activator of Nuclear Factor-kappa B; D048029:TNF Receptor-Associated Factor 6; D020928:Mitogen-Activated Protein Kinases; D051722:ADAM Proteins; D000072198:ADAM17 Protein