RA诱导SH—SY5Y细胞分化中NEF3基因表达的调控

目的 研究全反式维甲酸(RA)诱导神经母细胞瘤分化过程中NEF3表达的调控.方法 用RA诱导神经母细胞SH-SY5Y,用实时RT-PCR分析NEF3和BrgI的mRNA表达;并同时测定RGI蛋白的表达.共转染含有2.8kb NEF3上游序列的报告基因质粒、Brg1的表达质粒或者其负显性突变质粒,检测后两者对NEF3-CAT启动子活性的调控.结果 RA诱导12～24h,SH-SY5Y细胞中NEF3与Brg1几乎同时表达出现第1个高峰,之后表达量下降.共转染结果 显示Brgl可促进NEF3-CAT表达增加,而其突变型没有作用.结论 RA诱导初期,染色质重塑复合物ATP酶亚基Brg1可能导致NEF3上游调控序列所处的染色质结构发生改变,从而有利于NEF3基因的表达.

Regulation of the Expression of NEF3 in RA Induced Differentiation of SH-SYSY Neuroblastoma Cells

Objective To analyze the regulation of NEF3 expression in SH - SY5Y after RA induction. Methods The expression of NEF3 and Brg1 genes was individually detected by realtime RT - PCR and Western blot analyses. Then, the CAT reporter driven by 5' flanking region ofNEF3 gene (pBLCAT3 -2. 8k - NEF3) was co - transfccted with eukaryotic expression plasmids of pBJ5 - Brgl or pBJ5 - Brg1 - NTP into SH - SY5Y respectively. Promoter activity of the NEF3 gene was detected by competitive RT - PCR assay. Results Similar changes in the mRNA expression of NEF3 and Brg1 and that of the protein expression of Brgl were detected in Sh - SY5Y cells af-ter RA treatment for 12 -24h. Wild type Brgl can promote the expression of pBLCAT3 -2.8k -NEF3 promoter after RA induction, but not mutant Brg1. Conclusion Brgl helpes regulating the expression of NEF3 gene possibly via changes in chromatin conformation in the SH -SY5Y cells after RA induction.