Regulation of bone sialoprotein (BSP) gene transcription by lipopolysaccharide
Regulation of bone sialoprotein (BSP) gene transcription by lipopolysaccharide
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DOI：
10.1002/jcb.20628
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发表时间：
2006-02-01
影响因子：
4
通讯作者：
Ogata, Y
中科院分区：
文献类型：
--
作者：
Kato, N;Nakayama, Y;Ogata, Y
脂多糖（LPS）是牙周病炎症反应的主要介质，抑制骨形成并刺激骨吸收。为了确定抑制骨形成的分子机制，我们分析了LPS对BSP基因表达的影响。骨唾液蛋白（BSP）是一种矿化的组织特异性蛋白，似乎在骨的初始矿化中起作用。用LPS（1 μ g/ml）处理成骨细胞样ROS 17/2.8细胞12小时，导致BSP mRNA水平显著降低。在用LPS刺激前30分钟添加抗氧化剂N-乙酰半胱氨酸（NAC; 20 mM）减弱了BSP mRNA水平的抑制。瞬时转染分析，使用大鼠BSP基因启动子与荧光素酶报告基因连接的嵌合构建体，揭示LPS（1 μ g/ml）抑制转染入ROS 17/2.8细胞的荧光素酶构建体的表达，所述荧光素酶构建体包括BSP启动子核苷酸-108至+60。蛋白激酶A（PKA）抑制剂H89和酪氨酸激酶抑制剂除莠霉素A（HA）可抑制LPS的作用。在反向CCAAT盒（ATTGG; nts-50和-46）、cAMP反应元件（CRE; nts-75至-68）、FGF反应元件（FRE; nts-92至-85）和垂体特异性转录因子结合元件（Pit-1; nts-111至-105）中引入2 bp突变表明LPS效应由CRE和FRE介导。LPS处理后，FRE和3 '-FRE DNA-蛋白复合物减少，而CRE DNA-蛋白复合物无明显变化。因此，这些研究表明，LPS通过PKA和酪氨酸激酶依赖性途径抑制BSP基因转录，LPS的作用是通过近端BSP基因启动子中的CRE和FRE元件介导的。
Lipopolysaccharide (LPS) is a major mediator of inflammatory responses in periodontal disease that inhibits bone formation and stimulates bone resorption. To determine the molecular mechanisms involved in the suppression of bone formation, we have analyzed the effects of LPS on BSP gene expression. Bone sialoprotein (BSP) is a mineralized tissue-specific protein that appears to function in the initial mineralization of bone. Treatment of osteoblast like ROS 17/2.8 cells with LPS (1 mu g/ml) for 12 h caused a marked reduction in BSP mRNA levels. The addition of antioxidant N-acetylcysteine (NAC; 20 mM) 30 min prior to stimulation with LPS attenuated the inhibition of BSP mRNA levels. Transient transfection analyses, using chimeric constructs of the rat BSP gene promoter linked to a luciferase reporter gene, revealed that LPS (1 mu g/ml) suppressed expression of luciferase construct, encompassing BSP promoter nucleotides -108 to +60, transfected into ROS17/2.8 cells. The effects of LPS were inhibited by protein kinase A (PKA) inhibitor, H89 and the tyrosine kinase inhibitor, herbimycin A (HA). Introduction of 2 bp mutations in the inverted CCAAT box (ATTGG; nts -50 and -46), a cAMP response element (CRE; nts -75 to -68), a FGF response element (FRE; nts -92 to -85), and a pituitary specific transcription factor binding element (Pit-1; nts -111 to -105) showed that the LPS effects were mediated by the CRE and FRE. Whereas the FRE and 3'-FRE DNA-protein complexes were decreased by LPS, CRE DNA-protein complex did not change after LPS treatment. These studies, therefore, show that LPS suppresses BSP gene transcription through PKA and tyrosine kinase-dependent pathways and that the LPS effects are mediated through CRE and FRE elements in the proximal BSP gene promoter.