Insulin-like growth factor-II regulates bone sialoprotein gene transcription
Insulin-like growth factor-II regulates bone sialoprotein gene transcription
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DOI：
10.1007/s10266-015-0205-6
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发表时间：
2015-04
期刊：
影响因子：
2.5
通讯作者：
Jin Choe;Y. Sasaki;Liming Zhou;H. Takai;Y. Nakayama;Y. Ogata
中科院分区：
文献类型：
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作者：
Jin Choe;Y. Sasaki;Liming Zhou;H. Takai;Y. Nakayama;Y. Ogata
胰岛素样生长因子- i和-II （IGF-I和IGF-II）已在几种不同物种的骨提取物中被发现，而IGF-II是骨中储存最丰富的生长因子。骨涎蛋白（BSP）是一种与矿化结缔组织相关的非胶原细胞外基质糖蛋白。在本研究中，我们研究了IGF-II在大鼠成骨样ROS17/2.8细胞中对BSP转录的调控。IGF-II （50 ng/ml）刺激6 h后提高了BSP mRNA和蛋白水平，并增强了pLUC3（−116 ~ +60）、pLUC4（−425 ~ +60）、pLUC5（−801 ~ +60）和pLUC6（−938 ~ +60）的荧光素酶活性。酪氨酸激酶、细胞外信号调节激酶1/2和磷脂酰肌醇3激酶抑制剂可抑制IGF-II的作用，cAMP反应元件（CRE）、FGF2反应元件（FRE）和同源结构域蛋白结合位点（HOX）的2 bp突变可消除IGF-II的作用。凝胶转移实验结果显示，IGF-II （50 ng/ml）在3和6 h时增加了与CRE、FRE和HOX位点结合的核蛋白。CREB1、phospho-CREB1、c-Fos和c-Jun抗体破坏了CRE蛋白复合物的形成。Dlx5和Runx2抗体破坏了FRE -和hox -蛋白复合物的形成。因此，这些研究表明IGF-II通过靶向大鼠BSP基因近端启动子中的CRE、FRE和HOX元件来增加BSP的转录。此外，phospho-CREB1、c-Fos、c-Jun、Dlx5和Runx2转录因子似乎是IGF-II对BSP转录作用的关键调控因子。
Insulin-like growth factor-I and -II (IGF-I and IGF-II) have been found in bone extracts of several different species, and IGF-II is the most abundant growth factor stored in bone. Bone sialoprotein (BSP) is a noncollagenous extracellular matrix glycoprotein associated with mineralized connective tissues. In this study, we have investigated the regulation of BSP transcription by IGF-II in rat osteoblast-like ROS17/2.8 cells. IGF-II (50 ng/ml) increased BSP mRNA and protein levels after 6-h stimulation, and enhanced luciferase activities of the constructs pLUC3 (−116 to +60), pLUC4 (−425 to +60), pLUC5 (−801 to +60) and pLUC6 (−938 to +60). Effects of IGF-II were inhibited by tyrosine kinase, extracellular signal-regulated kinase1/2 and phosphatidylinositol 3-kinase inhibitors, and abrogated by 2-bp mutations in cAMP response element (CRE), FGF2 response element (FRE) and homeodomain protein-binding site (HOX). The results of gel shift assays showed that nuclear proteins binding to CRE, FRE and HOX sites were increased by IGF-II (50 ng/ml) at 3 and 6 h. CREB1, phospho-CREB1, c-Fos and c-Jun antibodies disrupted the formation of the CRE–protein complexes. Dlx5 and Runx2 antibodies disrupted the FRE– and HOX–protein complex formations. These studies therefore demonstrated that IGF-II increased BSP transcription by targeting CRE, FRE and HOX elements in the proximal promoter of the rat BSP gene. Moreover, phospho-CREB1, c-Fos, c-Jun, Dlx5 and Runx2 transcription factors appear to be key regulators of IGF-II effects on BSP transcription.