PI3KγmRNA 3′非翻译区可能存在基因表达负调控区

为探索人磷脂酰肌醇 3 激酶γ(phosphoinositide 3 kinasePI3Kγ)基因 3′端非翻译区内AU富含区是否在基因表达调控中起作用 ,首先通过生物信息学分析发现在其 3′端非翻译区 (UTR)内存在0 9kb的AU富含区 ,其中包括 4个AU富含元件 ,以及 1个与众多基因非翻译区高度同源的长 130个碱基的区域 .将AU富含区插入报告基因egfp的下游构建pcDNA3 egfp AUR表达载体 .将表达载体转导NIH 3T3,74 0 2及K5 6 2细胞 ,流式细胞检测egfp的表达情况 .PI3Kγ基因 3′非翻译区AU富含区可显著降低egfp的表达 2～ 3倍 (P <0 0 1) .利用放线菌素D阻断RNA转录后 ,Northern印迹分析结果显示egfp AURmRNA较egfpmRNA不稳定 .实验结果提示 ,PI3Kγ基因 3′非翻译区AU富含区内可能存在转录后水平的基因表达负调控区 ,该负调控区可在一定程度上加速mRNA的衰变

Probable Existence of a Negative Regulation Region Within the 3'UTR of Human Phosphoinositide 3-Kinaseγ mRNA

In order to study whether there is a functional regulation region within the 3′untranslational region (UTR) of human phosphoinositide 3 kinaseγ(PI3Kγ)mRNA, the 3′UTR was analyzed through bioinformatics. An AU rich region (AUR)of about 0 9 kb was found. Within the AU rich region there are 4 AU rich elements (AREs) and a region which is homologous with many other UTRs. The AU rich region was inserted into the 3′UTR of egfp reporter gene, and pcDNA3 egfp AUR expressing vector was constructed. The expression of egfp containing the 3′UTR of PI3Kγ was significantly reduced by 2～3 times in NIH 3T3, 7402 and K562 cells. After treated with actinomycin D (5 μg/ml) total RNA from stably transfected K562 cells were successively extracted with an interval of 2 hours, and the stability of egfp mRNA was analyzed through Northern blotting. The result showed that the AU rich region of PI3Kγ could, to some extent, accelerate the decay of egfp mRNA. These results indicate that there is probably a negative regulation region within the 3′AU rich region of PI3Kγ mRNA 3′UTR.