Department of Orthopedic Surgery, School of Medicine, Keio University, Tokyo 160-8582, Japan.
J Immunol. 2010 Mar 1;184(5):2442-8. doi: 10.4049/jimmunol.0901188. Epub 2010 Jan 29.
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.
破骨细胞生成是一个高度复杂的过程，涉及到在成骨细胞和破骨细胞前体中表达的多种膜结合蛋白。在过去的几年中，膜结合蛋白的蛋白水解裂解和胞外结构域的释放，也称为胞外结构域脱落，已成为一种重要的翻译后调节机制，用于修饰细胞表面蛋白的功能。与此观点一致，破骨细胞生成过程中涉及的几种膜结合分子，包括 CSF-1R 和核因子-κB 配体受体激活剂（RANKL），通过蛋白水解从细胞表面被裂解和释放。在这项研究中，我们研究了破骨细胞生成中最重要的分子之一 RANK 是否经历胞外结构域脱落。结果表明，RANK 以可溶性单体蛋白的形式释放，而 TNF-α 转换酶参与了这种活性。我们还鉴定了 RANK 近膜结构域中的潜在裂解位点，并发现 rRANKL 通过 TNFR 相关因子 6 和 MAPK 途径诱导 RAW264.7 样巨噬细胞系中的 RANK 脱落。此外，我们发现 TNF-α 转换酶缺陷型破骨细胞前体中的 RANKL 诱导的破骨细胞生成加速。这些观察结果表明，胞外结构域脱落可能参与 RANK 功能的调节，并为破骨细胞生成的机制提供新的见解。
