Wu Y, Mehew J W, Heckman C A, Arcinas M, Boxer L M
The Center for Molecular Biology in Medicine, Palo Alto Veterans Affairs Medical Center and the Department of Medicine, Stanford University School of Medicine, Stanford, CA 94305-5112, USA.
Oncogene. 2001 Jan 11;20(2):240-51. doi: 10.1038/sj.onc.1204067.
The p53 protein activates promoters containing p53 binding sites, and it represses other promoters. We examined the effect of p53 on bcl-2 expression in both the DHL-4 B cell line and the K562 erythroleukemia line. Transient transfection analyses revealed that wild-type p53 repressed the bcl-2 full-length promoter. The region of the bcl-2 promoter that was responsive to p53 was mapped to the bcl-2 P2 minimal promoter region, and we showed that p53 and the TATA binding protein bound to the bcl-2 TATA sequence. The TATA binding protein, p53, histone deacetylase-1 and mSin3a could be co-immunoprecipitated from K562 cell nuclear extract. The TATA binding protein and mSin3a could be recovered in a complex at the bcl-2 promoter TATA sequence, however, the formation of this complex was not dependent on the presence of p53. Treatment of K562 cells with the histone deacetylase inhibitor, trichostatin A, resulted in an increase in bcl-2 promoter activity whether p53 was present or not. Therefore, we demonstrated that p53 and the histone deacetylases repress the bcl-2 promoter independently. Similar results were obtained when endogenous bcl-2 mRNA or protein levels were measured in response to either p53 or trichostatin A, and p53 expression resulted in enhanced apoptosis. RNase protection assays demonstrated that transcription from the endogenous 3' bcl-2 promoter was decreased by p53. The regions of p53 that were required for repression of the bcl-2 promoter were defined. We conclude that the TATA sequence in the bcl-2 P2 minimal promoter is the target for repression by p53, and that the interaction between p53 and TBP is most likely responsible for the repression. Mutation of p53 may play a role in the up-regulation of bcl-2 expression in some B cell lymphomas.
p53蛋白可激活含有p53结合位点的启动子，并抑制其他启动子。我们研究了p53对DHL-4 B细胞系和K562红白血病细胞系中bcl-2表达的影响。瞬时转染分析显示，野生型p53可抑制bcl-2全长启动子。bcl-2启动子中对p53有反应的区域被定位到bcl-2 P2最小启动子区域，并且我们发现p53和TATA结合蛋白与bcl-2 TATA序列结合。TATA结合蛋白、p53、组蛋白去乙酰化酶-1和mSin3a可从K562细胞核提取物中进行共免疫沉淀。TATA结合蛋白和mSin3a可在bcl-2启动子TATA序列处的复合物中被回收，然而，该复合物的形成并不依赖于p53的存在。用组蛋白去乙酰化酶抑制剂曲古抑菌素A处理K562细胞，无论是否存在p53，bcl-2启动子活性均会增加。因此，我们证明p53和组蛋白去乙酰化酶可独立抑制bcl-2启动子。当检测内源性bcl-2 mRNA或蛋白水平对p53或曲古抑菌素A的反应时，获得了类似的结果，并且p53表达导致凋亡增强。核糖核酸酶保护分析表明，p53可降低内源性3' bcl-2启动子的转录。确定了抑制bcl-2启动子所需的p53区域。我们得出结论，bcl-2 P2最小启动子中的TATA序列是p53抑制的靶点，并且p53与TBP之间的相互作用最有可能是抑制的原因。p53突变可能在某些B细胞淋巴瘤中bcl-2表达上调中起作用。
