为了探讨人类UTROPHIN蛋白表达上调的可能因素, 文章利用P-match软件预测人类UTROPHIN基因启动子区域可能的调控元件, 应用CHIP和EMSA实验加以验证, 并利用RNA干扰和荧光定量PCR的方法分析人类EN1转录因子调节UTROPHIN表达的作用机制。经P-match软件预测, 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HeLa细胞中EN1基因沉默后, 可使UTROPHIN mRNA水平上调。以UTROPHIN蛋白为靶点可能为DMD(Duchenne muscular dystrophy)基因治疗提供一种新的策略。
To investigate possible factors up-regulating the expression of UTROPHIN, potential regulatory elements in the promoter of the human UTROPHIN was predicted by P-match software and verified by EMSA and ChIP. The mechanism of EN1 regulation of the human UTROPHIN expression was evaluated by RNA interference and real-time PCR analyses. Two potential EN1 binding sites in UTROPHIN promoter region were predicted by P-Match software but only the second site was verified to interact directly with EN1 by EMSA and ChIP. The results from RNA interference and real-time PCR showed that the mRNA level of UTROPHIN increased in HeLa cells after EN1 was knockdowned by siRNA. It indicated that EN1 might be a negative regulatory factor for UTROPHIN. Our study suggested that UTROPHIN might be a new target for DMD therapy.
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