Sp1特异性调控先天性心脏病相关基因钙调磷酸酶调节蛋白1基因亚型的转录

Sp1 specifically regulates the transcription of congenital heart disease associated gene RCAN1 isoform 1

目的探索先天性心脏病相关基因钙调磷酸酶调节蛋白1基因(RCAN1)两个亚型(RCAN1-1和RCAN1-4)组织特异性表达的机制。方法通过生物信息学预测RCAN1-1和RCAN1-4各自启动子区的转录因子结合位点,分析比对并筛选出具有差异的位点,通过定点突变及荧光素酶报告基因系统检测筛选出的位点活性。将鉴定的转录因子的抑制剂与RCAN1表达载体作用,分为实验组(抑制剂+)和对照组(抑制剂-),通过实时荧光定量PCR检测RCAN1亚型mRNA的表达量。结果在RCAN1-1启动子区-138GCCCGCCGCC-129处检测出一个新的、未报道过的具有活性的转录因子Sp1结合位点。实验组RCAN1-1 mRNA表达量低于对照组(P<0.001),而两组RCAN1-4 mRNA的表达量无差异。结论Sp1特异性调控RCAN1-1的转录,增强其表达,而对RCAN1-4无影响,提示其可作为将来基因治疗或检测的一个潜在靶点。

Objective To investigate the regulatory mechanism of regulator of calcineurin 1(RCAN1),which is associated with congenital heart disease and has a tissue-specific expression pattern.Methods The transcription factor binding sites were predicted and compared in promoter of RCAN1.The site directed mutation and luciferase assay were used to detect the activity of the binding site.Then the transcriptional regulation of RCAN1 by Sp1 was evaluated by real-time quantitative PCR under physiological condition(control group)or Sp1 inhibitor(experimental group).Results The Sp1 binding site-138GCCCGCCGCC-129 was predicated and identified.The RCAN1-1 mRNA was more decreased in experimental group than the control group(P<0.001).There was no significant difference in RCAN1-4 mRNA between the two groups.Conclusions Sp1 specifically regulates the transcription of RCAN1-1 isoform,which demonstrates that Sp1 is a potential targete molecule for genetic testing and clinical intervention for congenital heart disease.