Shao J, Fujiwara T, Kadowaki Y, Fukazawa T, Waku T, Itoshima T, Yamatsuji T, Nishizaki M, Roth J A, Tanaka N
First Department of Surgery, Okayama University Medical School, Japan.
Oncogene. 2000 Feb 10;19(6):726-36. doi: 10.1038/sj.onc.1203383.
The tumor suppressor gene p53 is a potent transcriptional regulator of genes which are involved in many cellular activities including cell cycle arrest, apoptosis, and angiogenesis. Recent studies have demonstrated that the activation of the transcriptional factor nuclear factor kappaB (NF-kappaB) plays an essential role in preventing apoptotic cell death. In this study, to better understand the mechanism responsible for the p53-mediated apoptosis, the effect of wild-type p53 (wt-p53) gene transfer on nuclear expression of NF-kappaB was determined in human colon cancer cell lines. A Western blot analysis of nuclear extracts demonstrated that NF-kappaB protein levels in the nuclei were suppressed by the transient expression of the wt-p53 in a dose-dependent manner. Transduced wt-p53 expression increased the cytoplasmic expression of I kappaB alpha as well as its binding ability to NF-kappaB, thus markedly reducing the amount of NF-kappaB that translocated to the nucleus. The decrease in nuclear NF-kappaB protein correlated with the decreased NF-kappaB constitutive activity measured by electrophoretic mobility shift assay. Furthermore, parental cells transfected with NF-kappaB were better protected from cell death induced by the wt-p53 gene transfer. We also found that the wt-p53 gene transfer was synergistic with aspirin (acetylsalicylic acid) in inhibiting NF-kappaB constitutive activity, resulting in enhanced apoptotic cell death. These results suggest that the inhibition of NF-kappaB activity is a plausible mechanism for apoptosis induced by the wt-p53 gene transfer in human colon cancer cells and that anti-NF-kappaB reagent aspirin could make these cells more susceptible to apoptosis.
肿瘤抑制基因p53是一种强大的转录调节因子，可调节许多参与细胞活动的基因，这些活动包括细胞周期停滞、细胞凋亡和血管生成。最近的研究表明，转录因子核因子κB（NF-κB）的激活在防止凋亡性细胞死亡中起着至关重要的作用。在本研究中，为了更好地理解p53介导的细胞凋亡机制，我们在人结肠癌细胞系中测定了野生型p53（wt-p53）基因转移对NF-κB核表达的影响。对细胞核提取物进行的蛋白质免疫印迹分析表明，wt-p53的瞬时表达以剂量依赖的方式抑制了细胞核中NF-κB蛋白的水平。转导的wt-p53表达增加了IκBα的细胞质表达及其与NF-κB的结合能力，从而显著减少了易位至细胞核的NF-κB的量。细胞核中NF-κB蛋白的减少与通过电泳迁移率变动分析测定的NF-κB组成型活性的降低相关。此外，用NF-κB转染的亲代细胞对wt-p53基因转移诱导的细胞死亡具有更好的保护作用。我们还发现，wt-p53基因转移与阿司匹林（乙酰水杨酸）在抑制NF-κB组成型活性方面具有协同作用，从而导致凋亡性细胞死亡增加。这些结果表明，抑制NF-κB活性是wt-p53基因转移诱导人结肠癌细胞凋亡的一种合理机制，并且抗NF-κB试剂阿司匹林可使这些细胞更容易发生凋亡。
