【摘要】 目的探讨人类横纹肌肉瘤细胞中Utrophin蛋白表达上调的可能因素。方法查阅文献,检索人类Utrophin基因启动子区域可能的调控元件,应用染色质免疫沉淀(CHIP)和凝胶阻滞实验(EMSA)加以验证。并利用RNA干扰和荧光定量聚合酶链反应(PCR)的方法分析人类EN1转录因子调节Utrophin表达的作用机制。结果经查阅文献获知,Utrophin基因启动子区域有2个转录因子EN1的可能结合位点,经CHIP和EMSA实验证实位点2是真正结合位点。通过RNA干扰实验,发现在人横纹肌肉瘤细胞中EN1基因沉默后,可使Utrophin mRNA水平上调。结论如果能进一步验证其蛋白表达水平亦升高,将可能为假肥大性肌营养不良基因治疗提供新的靶点。更多还原

【Abstract】 Objective To investigate the factor of up-regulating the expression of utrophin protein in human rhabdomyosarcoma cells.Methods The regulatory element in the promoter region of the human utrophin was searched through documents and further verified by CHIP and EMSA experiment.Meanwhile,the mechanism of EN1 regulation of the human utrophin was evaluated by RNA interference and real-time PCR analysis.Results Two potential EN1 binding sites in utrophin promoter region were found by searching the documents,but only the 2nd site was verified to interact directly with EN1 by EMSA and CHIP.The results from RNA interference,real-time PCR analysis suggested that after EN1 was knocked down by siRNA,the expression of utrophin mRNA was increased in rhabdomyosarcoma cells.Conclusion It indicated that EN1 might be the negative regulatory element in the upstream of utrophin.Our study suggested that enhancement of utrophin promoter activation may be as a therapeutic strategy for Duchenne muscular dystrophy.更多还原