Proteomic analysis of lysosomal acid hydrolases secreted by osteoclasts - Implications for lytic enzyme transport and bone metabolism
Proteomic analysis of lysosomal acid hydrolases secreted by osteoclasts - Implications for lytic enzyme transport and bone metabolism
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DOI：
10.1074/mcp.m500291-mcp200
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发表时间：
2006-01-01
影响因子：
7
通讯作者：
Hoflack, B
中科院分区：
文献类型：
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作者：
Czupalla, C;Mansukoski, H;Hoflack, B
破骨细胞是骨消化细胞，是极化细胞，它们分泌酸性水解酶到发生骨降解的再吸收腔隙中。这一过程背后的分子机制尚不清楚。为了分析破骨细胞分泌的酸性水解酶的性质，我们使用了小鼠骨髓 Raw 264.7 细胞系，该细胞系在 NF-κ B 配体受体激活剂存在的情况下在体外分化为成熟的破骨细胞。分化后，我们观察到含甘露糖 6-磷酸的酸性水解酶的分泌强烈增加。对甘露糖 6-磷酸受体亲和柱上捕获的分泌蛋白进行蛋白质组学分析，发现有 58 种不同的蛋白质属于多个酸性水解酶家族，其中 16 种明显参与骨稳态。此外，这些酸性水解酶作为前蛋白被分泌。大多数已鉴定的酸性水解酶的表达在破骨细胞生成过程中没有变化。因此，我们的数据强烈支持这样的观点，即破骨细胞对酸性水解酶的极化分泌是由于沿溶酶体途径的膜运输的关键步骤的重组造成的，而不是由于溶酶体与面向吸收腔隙的膜的融合造成的。
Osteoclasts, the bone-digesting cells, are polarized cells that secrete acid hydrolases into a resorption lacuna where bone degradation takes place. The molecular mechanisms underlying this process are poorly understood. To analyze the nature of acid hydrolases secreted by osteoclasts, we used the mouse myeloid Raw 264.7 cell line that differentiates in vitro into mature osteoclasts in the presence of the receptor activator of NF-kappa B ligand. Upon differentiation, we observed a strong increase in the secretion of mannose 6-phosphate-containing acid hydrolases. A proteomic analysis of the secreted proteins captured on a mannose 6-phosphate receptor affinity column revealed 58 different proteins belonging to several families of acid hydrolases of which 16 are clearly involved in bone homeostasis. Moreover these acid hydrolases were secreted as proproteins. The expression of most of the identified acid hydrolases is unchanged during osteoclastogenesis. Thus, our data strongly support the notion that the polarized secretion of acid hydrolases by osteoclasts results from a reorganization of key steps of membrane traffic along the lysosomal pathway rather than from a fusion of lysosomes with the membrane facing the resorption lacuna.