Identification of a homeodomain binding element in the bone sialoprotein gene promoter that is required for its osteoblast-selective expression
Identification of a homeodomain binding element in the bone sialoprotein gene promoter that is required for its osteoblast-selective expression
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DOI：
10.1074/jbc.275.18.13907
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发表时间：
2000-05-05
影响因子：
4.8
通讯作者：
Franceschi, RT
中科院分区：
文献类型：
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作者：
Benson, MD;Bargeon, JL;Franceschi, RT
骨唾液蛋白是一种70 kda的细胞外基质成分，与生物矿化密切相关，但Bsp基因中限制其在矿化细胞中表达的顺式作用元件仍未被表征。为了确定这些元件，我们分析了小鼠启动子的2472碱基对片段，该片段指导荧光素酶报告基因的成骨选择性表达，并发现相对于转录起点的-338和-178之间的区域对其成骨选择性活性至关重要。我们在这个区域内发现了一个元件，该元件与成骨细胞核提取物中的蛋白质复合物结合，并且是其转录活性所必需的。引入一个突变，破坏该序列内的同源结构域结合位点，消除其体外结合和几乎所有2472碱基对启动子的成骨选择性活性。我们进一步发现，能够调控成骨特异性骨钙素启动子的Dlx5同源蛋白在COS7细胞中过表达时，可以结合该元件并刺激其增强子活性。这些数据首次描述了骨唾液蛋白启动子内的成骨细胞特异性元件，并证明了其受已知参与骨骼形成的因子家族成员的调节。
Bone sialoprotein is a 70-kDa extracellular matrix component that is intimately associated with biomineralization, yet the cis-acting elements of the Bsp gene that restrict its expression to mineralizing cells remain uncharacterized. To identify such elements, we analyzed a 2472-base pair fragment of the murine promoter that directs osteoblast-selective expression of a luciferase reporter gene and found that the region between -338 and -178 relative to the transcriptional start is crucial for its osteoblast-selective activity. We identified an element within this region that binds a protein complex in the nuclear extracts of osteoblastic cells and is required for its transcriptional activity. Introduction of a mutation that disrupts a homeodomain binding site within this sequence eliminates both its in vitro binding and nearly all of the osteoblastic-selective activity of the 2472-base pair promoter. We further found that the Dlx5 homeoprotein, which is able to regulate the osteoblast-specific osteocalcin promoter, can bind this element and stimulate its enhancer activity when overexpressed in COS7 cells. These data represent the first description of an osteoblast-specific element within the bone sialoprotein promoter and demonstrate its regulation by a member of a family of factors known to be involved in skeletogenesis.