转录因子Runx2调控前成骨细胞外基质磷酸化糖蛋白基因启动子的表达
发布时间：2018-11-11 12:28
【摘要】：背景:细胞外基质磷酸化糖蛋白基因在骨的矿化和吸收、成骨细胞与破骨细胞的平衡中起重要的作用,研究细胞外基质磷酸化糖蛋白的功能及其调控机制可为骨质疏松症的治疗提供新的思路。目的:分析转录因子Runx2在小鼠前成骨细胞中对细胞外基质磷酸化糖蛋白基因启动子的调控作用,进而初步研究转录因子Runx2在骨形成发育过程中的作用。方法:首先根据Genbank中Runx2的基因序列构建Runx2真核表达载体;然后利用双荧光素酶基因检测报告系统分析Runx2对不同长度的细胞外基质磷酸化糖蛋白基因启动子转录活性的影响,以确定Runx2有明显作用的启动子区段,分析3种MAPK信号通路抑制剂调控Runx2对细胞外基质磷酸化糖蛋白基因启动子转录活性的影响;最后利用定时定量RT-PCR法分析Runx2对细胞外基质磷酸化糖蛋白基因启动子表达活性的影响。结果与结论:成功构建Runx2真核表达载体;双荧光素酶基因检测报告系统分析显示Runx2能够上调细胞外基质磷酸化糖蛋白基因启动子在前成骨细胞中的转录活性,在(-300-+66)366 bp片段区域内上调效果较为显著,Runx2可通过激活MEK激酶MAPK通路上调细胞外基质磷酸化糖蛋白启动子活性;定时定量RT-PCR法检测再次验证Runx2上调细胞外基质磷酸化糖蛋白基因启动子的表达水平。提示转录因子Runx2可以通过MEK激酶MAPK信号通路对细胞外基质磷酸化糖蛋白基因表达进行调节,为探讨细胞外基质磷酸化糖蛋白在骨形成发育过程中的意义奠定基础。
[Abstract]:Background: extracellular matrix phosphorylated glycoprotein gene plays an important role in the mineralization and absorption of bone and the balance between osteoblasts and osteoclasts. Studying the function of extracellular matrix phosphorylated glycoprotein and its regulatory mechanism may provide new ideas for the treatment of osteoporosis. Aim: to investigate the role of transcription factor Runx2 in the regulation of extracellular matrix phosphorylated glycoprotein gene promoter in mouse preosteoblasts, and to investigate the role of transcription factor Runx2 in bone formation and development. Methods: firstly, the eukaryotic expression vector of Runx2 was constructed according to the gene sequence of Runx2 in Genbank. Then the effects of Runx2 on the transcriptional activity of different length extracellular matrix phosphorylated glycoprotein gene promoter were analyzed by double luciferase gene detection report system to determine the promoter region of Runx2. The effects of three MAPK signal pathway inhibitors on the transcriptional activity of extracellular matrix phosphorylated glycoprotein gene promoter were analyzed. Finally, the effect of Runx2 on the expression of extracellular matrix phosphorylated glycoprotein gene promoter was analyzed by timed quantitative RT-PCR. Results & conclusion: Runx2 eukaryotic expression vector was successfully constructed. Double luciferase gene detection system analysis showed that Runx2 could up-regulate the transcriptional activity of extracellular matrix phosphorylated glycoprotein gene promoter in preosteoblasts, and significantly increased the activity in (-300-66) 366 bp region. Runx2 upregulated the activity of extracellular matrix phosphorylated glycoprotein promoter by activating MEK kinase MAPK pathway. The expression of extracellular matrix phosphorylated glycoprotein gene promoter was up-regulated by Runx2. It is suggested that transcription factor Runx2 can regulate the expression of extracellular matrix phosphorylated glycoprotein gene through MEK kinase MAPK signaling pathway, which lays a foundation for the study of the significance of extracellular matrix phosphorylated glycoprotein in bone formation and development.
【作者单位】： 潍坊医学院口腔医学院;潍坊医学院附属医院;潍坊医学院;
【基金】：国家自然科学基金(81441107) 山东省自然科学基金(ZR2012HQ036) 潍坊医学院大学生科技创新基金项目(KX2013011)~~
【分类号】：R318.08
[Abstract]:Background: extracellular matrix phosphorylated glycoprotein gene plays an important role in the mineralization and absorption of bone and the balance between osteoblasts and osteoclasts. Studying the function of extracellular matrix phosphorylated glycoprotein and its regulatory mechanism may provide new ideas for the treatment of osteoporosis. Aim: to investigate the role of transcription factor Runx2 in the regulation of extracellular matrix phosphorylated glycoprotein gene promoter in mouse preosteoblasts, and to investigate the role of transcription factor Runx2 in bone formation and development. Methods: firstly, the eukaryotic expression vector of Runx2 was constructed according to the gene sequence of Runx2 in Genbank. Then the effects of Runx2 on the transcriptional activity of different length extracellular matrix phosphorylated glycoprotein gene promoter were analyzed by double luciferase gene detection report system to determine the promoter region of Runx2. The effects of three MAPK signal pathway inhibitors on the transcriptional activity of extracellular matrix phosphorylated glycoprotein gene promoter were analyzed. Finally, the effect of Runx2 on the expression of extracellular matrix phosphorylated glycoprotein gene promoter was analyzed by timed quantitative RT-PCR. Results & conclusion: Runx2 eukaryotic expression vector was successfully constructed. Double luciferase gene detection system analysis showed that Runx2 could up-regulate the transcriptional activity of extracellular matrix phosphorylated glycoprotein gene promoter in preosteoblasts, and significantly increased the activity in (-300-66) 366 bp region. Runx2 upregulated the activity of extracellular matrix phosphorylated glycoprotein promoter by activating MEK kinase MAPK pathway. The expression of extracellular matrix phosphorylated glycoprotein gene promoter was up-regulated by Runx2. It is suggested that transcription factor Runx2 can regulate the expression of extracellular matrix phosphorylated glycoprotein gene through MEK kinase MAPK signaling pathway, which lays a foundation for the study of the significance of extracellular matrix phosphorylated glycoprotein in bone formation and development.
【作者单位】： 潍坊医学院口腔医学院;潍坊医学院附属医院;潍坊医学院;
【基金】：国家自然科学基金(81441107) 山东省自然科学基金(ZR2012HQ036) 潍坊医学院大学生科技创新基金项目(KX2013011)~~
【分类号】：R318.08
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