三乙烯四胺对c-MYC启动子的调节作用01.doc

三乙烯四胺对c-MYC启动子的调节作用 邓小红 刘建辉* 郑旭煦 陈刚 郭丽霞 重庆工商大学药物化学与化学生物学研究中心，重庆，400067 摘要 目的 探讨三乙烯四胺 (triethylene tetramine, TETA)对c-MYC启动子的调节作用。方法 构建c-MYC启动子的荧光报告质粒及其突变体，经过序列测定后，转染HEK293细胞24 h后，以终浓度为0 (mol/L, 0.1 (mol/L, 1 (mol/L, 10 (mol/L, 100 (mol/L的TETA处理，测定其启动子的转录活性，计算TETA对其转录活性抑制率。结果 成功构建c-MYC启动子荧光报告质粒PGL3-Basic/c-MYC NHE III1 promoter及其突变体pGL3-Basci/c-MYC NHEIII1 promoter mutant。将二者分别转染细胞后发现，TETA可以剂量依赖的抑制c-MYC启动子的转录活性，而对突变体的转录活性抑制作用明显下降。结论 TETA能通过c-MYC启动子上的超敏元件对其转录活性具有负调节作用。 关键词： TETA；c-MYC启动子；G四链体；转录活性 基金项目：国家自然基金、教育部新世纪优秀人才计划 （NCET-07-0913）以及重庆市科委重点基础项目（CSTC, 2005BA5023）的资助。 作者简介:邓小红，女，在读博士　*通讯作者: 刘建辉，男，教授，硕士研究生导师　　Tel: (023)　Email: jhliu@ctbu.edu.cnTETA regulates the transcription of c-MYC promoter by enhancing the stability of G-quadruplex DENG Xiao-hong, LIU Jian-hui*, ZHENG Xu-xu, CHEN Gang, GUO Li-xia (Research Center of Pharmaceutical Chemistry Chemical Biology, Chongqing Technology and Business University, Chongqing, 400067, China) ABSTRACT: OBJECTIVE To study the effect of triethylene tetramine (TETA) on the transcription of c-MYC promoter. METHODS After the wild and mutant reporter gene plasmids containing the c-MYC NHE III1 sequence were constructed, the two plasmid were transfected into HEK 293 cells. The transfected cells were replated into 96 wells plate, and treated with different concentrations of TETA (0.0 (mol/L, 0.1 (mol/L, 1 (mol/L, 10 (mol/L, 100 (mol/L) for about 6-8h, the luciferase activity was determined with its substrate BrightGlo. The inhibiting rate of TETA on the reporter gene were calculated by the luciferase activity. RESULTS The luciferase report gene plasmids including pGL3-Basic/c-MYC NHE III1 promoter and its mutant were constructed successfully. And TETA could inhibit the transcription activity of wild reporter gene in a dose-dependent manner, but for the mutated gene, the inhibiting rate was decreased significantly. CONCLUSION TTETA has negative regulatory effect on c-MYC promoter through nucl