YY1对胶质瘤细胞迁移能力的影响及其分子机制

Effect of Yin Yang 1 on the migration ability of gliomacells and its molecular mechanism

目的探讨转录因子阴阳1(YY1)对胶质瘤细胞迁移的影响及其分子机制。方法体外培养人胶质瘤细胞U251和U87,对细胞过表达或敲低YY1后,通过Transwell实验、细胞划痕实验、克隆形成实验检测过表达或敲低YY1对胶质瘤细胞U251和U87迁移的影响;通过基因转录调控数据库(GTRD)预测YY1下游靶基因,应用实时荧光定量PCR(qRT-PCR)和蛋白印迹实验检测YY1下游靶基因基质金属蛋白酶15(MMP15)mRNA及其蛋白的表达情况;利用双荧光素酶报告实验检测YY1与MMP15启动子的结合区域。通过Transwell实验、细胞划痕实验、克隆形成实验检测过表达HA-YY1的同时敲低MMP15对胶质瘤细胞U251和U87迁移的影响。结果Transwell实验结果表明,与未过表达HA-YY1的对照组相比,过表达HA-YY1组U251和U87细胞的迁移能力显著提高(t=13.300、5.847,P<0.05);划痕实验结果显示,过表达HA-YY1组U251和U87细胞的划痕间距较对照组明显变小(t=9.698、7.215,P<0.05);克隆形成实验结果显示,过表达HA-YY1组U251和U87细胞的克隆形成能力较对照组显著提高(t=5.871、5.095,P<0.05);双荧光素酶报告实验显示,YY1能够直接结合到MMP15的启动子区域,并促进MMP15的表达;而敲低MMP15可以抑制由于YY1过表达而促进的细胞迁移。结论YY1通过转录激活MMP15促进胶质瘤细胞迁移。

Objective To investigate the effect of the transcription factor Yin Yang 1(YY1)on the migration of glioma cells and its molecular mechanism.Methods Human glioma U251 and U87 cells were cultured in vitro,and after the overexpression or knockdown of YY1,Transwell assay,cell scratch assay,and colony formation assay were used to observe the effect of YY1 overexpression or knockdown on the migration of glioma U251 and U87 cells.The Gene Transcription Regulation Database was used to predict the downstream targetgenes of YY1,and quantitative real-time PCR and Western blot were used to measure the mRNA and protein expression levels of matrix metalloproteinase 15(MMP15),a downstream target gene of YY1.Dual-luciferase reporter assay was used to investigate the binding region between YY1 and MMP15 promoter.Transwell assay,cell scratch assay,and colony formation assay were used to investigate the effect of HA-YY1 overexpression and MMP15 knockdown on the migration of glioma U251 and U87 cells.Results Transwell assay showed that compared with the control group without HA-YY1 overexpression,the HA-YY1 overexpression group had a significant increase in the migration ability of U251 and U87 cells(t=13.300,5.847;P<0.05).Cell scratch assay showed that the HA-YY1 overexpression group had a significant reduction in scratch spacing(t=9.698,7.215;P<0.05).Colony formation assay showed significant increases in the colony formation ability of U251 and U87 cells with HA-YY1 overexpression(t=5.871,5.095;P<0.05).Dual-luciferase reporter assay showed that YY1 could directly bind to the promoter region of MMP15 and promote the expression of MMP15,while knockdown of MMP15 could inhibit cell migration promoted by YY1 overexpression.Conclusion YY1 promotes the migration of glioma cells through transcriptional activation of MMP15.