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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 effect of AML1-ETO fusion gene on the transcriptional activity of p21WAF1 / CIP1 promoter and to explore the mechanism of AML1-ETO promoting leukemia. Methods The reporter plasmid of p21WAF1 / CIP1 promoter was constructed and co-transfected with the expression vector of AML1-ETO, AML1b and AML1a to determine the luciferase activity in CV-1 cells of African green monkey kidney cell line. The AML1-ETO, AML1b And AML1a on p21WAF1 / CIP1 gene promoter transcriptional activity. Results The transcription of p21WAF1 / CIP1 promoter was significantly inhibited by AML1-ETO in CV-1 cells. When the dose of pCMV5-AML1-ETO was 1000 ng, the transcriptional activity of p21 WAF1 / CIP1 promoter was decreased to The effect of AML1b and AML1a on the promoter activity of p21 WAF1 / CIP1 gene was not obvious. At the dose of 1000 ng, p21WAF1 / The CIP1 promoter transcriptional activity decreased to (61 ± 16)% and (59 ± 16)% of the control group, respectively. CONCLUSIONS: After AML1 is fused with ETO, the product of transcriptional repression of AML1 and AML1b can be more efficiently recruited by ETO protein because of its ability to recruit transcriptional repressor complexes. The effect of exogenous AML1-ETO on p21WAF1 / CIP1 May also be related to cell lines.