AML1-ETO对p21WAF1/CIP1启动子转录活性作用的研究

Effects of AML1-ETO on transcription activity of p21WAF1/CIP1 gene promoter

目的观察 AML1-ETO 融合基因对 p21WAF1/CIP1 基因启动子转录活性的影响,探讨AML1-ETO 促进白血病发生的机制。方法构建 p21WAF1/CIP1 基因启动子的报告质粒,与 AML1-ETO、AML1b 和 AML1a 的表达质粒共转染非洲绿猴肾细胞系 CV-1细胞,测定荧光素酶的活性,分析AML1-ETO、AML1b 和 AML1a 对 p21WAF1/CIP1基因启动子转录活性的影响。结果在 CV-1细胞中,AML1-ETO 对 p21WAF1/CIP1基因启动子的转录具有明显的抑制作用,在 pCMV5-AML1-ETO 的剂量为1000 ng 时,p21 WAF1/CIP1启动子的转录活性下降为对照组的(19±4)%,这种作用具有序列特异性和剂量依赖性;AML1b 和 AML1a 对 p21 WAF1/CIP1基因启动子转录活性的抑制作用不明显,在剂量为1000 ng 时,p21WAF1/CIP1启动子的转录活性分别下降为对照组的(61±16)%和(59±16)%。结论 AML1在与 ETO 形成融合基因后,其产物由于 ETO 蛋白能够更有效地募集转录共抑制复合物,其转录抑制活性要比 AML1a 和 AML1b 更强;外源的 AML1-ETO 对 p21WAF1/CIP1的作用可能也与细胞系有关。

Objective To observe the effects of AML1-ETO fusion gene on the transcription activity of p21WAF1/CIP1 gene. And to explore the enhancement of leukemia pathogenesis of AML1-ETO. Methods The luciferase reporter plasmids of 1321WAF1/CIP1 gene promoter were constructed, and co-transfected into CV-1 cells with AML1-ETO,AML1 b and AML1 a expression plasmids. The transactivity of p21WAF1/CIP1 gene promoter was assayed by luminometer. Results AML1-ETO exhibited a distinct inhibition activity of p21WAF1/CIP1 gene promoter with a sequence-specificity and dosage-dependant manner. The transactivity of p21WAF1/CIP1 gene promoter decreased to （19 ±4）% compared to control group, when 1000 ng pCMV5- AML1-ETO plasmid was used. AMLlb and AMLla showed less inhibition activity. The transactivity of p21WAF1/CIP1 gene promoter decreased to （61 ± 16）% and （59 ± 16）% compared to control group, respectively, when 1000 ng plasmid was used. Conclusion AML1-ETO exhibits more inhibition activity of p21WAF1/CIP｜ gene promoter than AML1 b and AML1 a, results from recruiting transcription co-repression complex efficiently by ETO. Based on previous researches, the effects of exogenous AML1-ETO on p21WAF1/CIP1 gene promote may be dependent on the type of cell lines.