Park Seyeon, Hahm Eun-Ryeong, Lee Dug-Keun, Yang Chul-Hak
School of Chemistry and Molecular Engineering, Seoul National University, Seoul 151-742, Korea.
J Cell Biochem. 2004 Apr 1;91(5):973-86. doi: 10.1002/jcb.10768.
Transcriptional activation of AP-1 is intricately involved in cell proliferation and transformation. The natural product, nordihydroguaiaretic acid (NDGA) shows an inhibitory effect on the binding of jun/AP-1 protein to the AP-1 site in 12-O-tetradecanoylphorbol-13-acetate (TPA)-stimulated HL60 cells. The NDGA inhibits the auto-regulated de novo synthesis of c-jun mRNA in TPA-stimulated HL60 cells. Our data also determine that this compound induces proliferation inhibition and apoptosis in human leukemia HL60 cells. To obtain information on the functional role of the AP-1 inhibition by NDGA in apoptosis signaling, the effects of pharmacological inhibition of AP-1 binding on c-myc, p53, and bax protein level were determined. Our results indicate that treatment of cells with NDGA enhances c-myc, p53, and bax protein levels. To rule out the possibility that NDGA will induce apoptosis because of the effects on proteins other than AP-1, we investigated the effect of another AP-1 inhibitor, SP600125, which is specific to Jun-N-terminal kinase. SP600125 decreased not only the phosphorylation level of jun protein but also AP-1/DNA binding activity. Also, apoptosis was observed to be induced by SP600125, concomitant with the increase in c-myc, p53, and bax protein level. In addition, apoptosis induced by both AP-1 inhibitors was accompanied by the activation of a downstream apoptotic cascade such as caspase 9, caspase 3, and poly[ADP-ribose]polymerase (PARP). When the cells were treated with NDGA or SP600125 in the presence of antisense c-myc oligonucleotides, apoptosis was not observed and an increase of c-myc, p53, and bax proteins was not manifested. All these results show that the inhibition of the transcription factor AP-1 action is related with either the drug-induced apoptosis or the drug toxicity of the HL60 cells. The apoptosis induced by AP-1 inhibition may be dependent on c-myc protein levels suggesting that the c-myc protein induces apoptosis at a low level of AP-1 binding activity. Altogether, our findings suggest that the presence of the AP-1 signal acts as a survival factor that determines the outcome of myc-induced proliferation or apoptosis.
AP-1的转录激活与细胞增殖和转化密切相关。天然产物去甲二氢愈创木酸（NDGA）对12-O-十四酰佛波醇-13-乙酸酯（TPA）刺激的HL60细胞中jun/AP-1蛋白与AP-1位点的结合具有抑制作用。NDGA抑制TPA刺激的HL60细胞中c-jun mRNA的自调控从头合成。我们的数据还表明，该化合物可诱导人白血病HL60细胞的增殖抑制和凋亡。为了获得关于NDGA抑制AP-1在凋亡信号传导中的功能作用的信息，我们确定了AP-1结合的药理学抑制对c-myc、p53和bax蛋白水平的影响。我们的结果表明，用NDGA处理细胞可提高c-myc、p53和bax蛋白水平。为了排除NDGA因对AP-1以外的蛋白质产生影响而诱导凋亡的可能性，我们研究了另一种AP-1抑制剂SP600125的作用，它对Jun N端激酶具有特异性。SP600125不仅降低了jun蛋白的磷酸化水平，还降低了AP-1/DNA结合活性。此外，观察到SP600125诱导了凋亡，同时c-myc、p53和bax蛋白水平增加。此外，两种AP-1抑制剂诱导的凋亡均伴随着下游凋亡级联反应的激活，如半胱天冬酶9、半胱天冬酶3和聚[ADP-核糖]聚合酶（PARP）。当在反义c-myc寡核苷酸存在的情况下用NDGA或SP600125处理细胞时，未观察到凋亡，且c-myc、p53和bax蛋白也未增加。所有这些结果表明，转录因子AP-1作用的抑制与HL60细胞的药物诱导凋亡或药物毒性有关。AP-1抑制诱导的凋亡可能依赖于c-myc蛋白水平，这表明c-myc蛋白在低水平的AP-1结合活性下诱导凋亡。总之，我们的研究结果表明，AP-1信号的存在作为一种生存因子，决定了myc诱导的增殖或凋亡的结果。
