Kang K H, Kim W H, Choi K H
Department of Life Science, College of Natural Sciences, Chung-Ang University, Seoul, 156-756, Korea.
Exp Cell Res. 1999 Dec 15;253(2):403-12. doi: 10.1006/excr.1999.4644.
p21, a potent cyclin-dependent kinase inhibitor, has been known to induce cell cycle arrest in response to DNA-damaging agents. Although p21 has been reported to play an important role in the regulation of apoptosis, the postulated role for p21 in apoptosis is still controversial. Previously, we reported that p21 was induced in a p53-independent manner during ceramide-induced apoptosis in human hepatocarcinoma cell lines. In the present study, we investigated the precise role of p21 in ceramide-induced apoptosis in human hepatocarcinoma cells by using a tetracycline-inducible expression system. Overexpression of p21 by itself did not induce apoptosis in p53-deficient Hep3B cells. However, Hep3B/p21 cells were more sensitive to ceramide-induced apoptosis. In these cells, p21 overexpression did not result in G1 arrest. The expression level of Bax was increased in Hep3B/p21 cells treated with ceramide and its expression was more accelerated under the p21-overexpressed condition compared to that of the p21-repressed condition. Overexpression of Bax induced apoptosis in Hep3B cells. On the other hand, the levels of p21 and Bax protein were increased by ceramide in another hepatocarcinoma cell line, SK-Hep-1, while the Bcl-2 protein level was not changed. Overexpression of Bcl-2 not only suppressed apoptosis but also completely prevented induction of p21 and Bax caused by ceramide in SK-Hep-1 cells. Furthermore, overexpression of p21 antagonized the death-protective function of Bcl-2 and upregulated expression of Bax protein. These results suggest that p21 promotes ceramide-induced apoptosis by enhancing the expression of Bax, thereby modulating the molecular ratio of Bcl-2:Bax in human hepatocarcinoma cells.
p21是一种有效的细胞周期蛋白依赖性激酶抑制剂，已知其可响应DNA损伤剂诱导细胞周期停滞。尽管有报道称p21在细胞凋亡调控中发挥重要作用，但p21在细胞凋亡中假定的作用仍存在争议。此前，我们报道在人肝癌细胞系中，神经酰胺诱导的细胞凋亡过程中p21以不依赖p53的方式被诱导。在本研究中，我们通过使用四环素诱导表达系统研究了p21在人肝癌细胞中神经酰胺诱导的细胞凋亡中的精确作用。p21自身过表达并未在p53缺陷的Hep3B细胞中诱导细胞凋亡。然而，Hep3B/p21细胞对神经酰胺诱导的细胞凋亡更敏感。在这些细胞中，p21过表达并未导致G1期停滞。用神经酰胺处理的Hep3B/p21细胞中Bax的表达水平升高，且与p21抑制状态相比，在p21过表达状态下其表达加速更明显。Bax过表达在Hep3B细胞中诱导细胞凋亡。另一方面，在另一个肝癌细胞系SK-Hep-1中，神经酰胺使p21和Bax蛋白水平升高，而Bcl-2蛋白水平未改变。Bcl-2过表达不仅抑制细胞凋亡，还完全阻止了SK-Hep-1细胞中神经酰胺引起的p21和Bax的诱导。此外，p21过表达拮抗了Bcl-2的抗死亡功能并上调了Bax蛋白的表达。这些结果表明，p21通过增强Bax的表达促进神经酰胺诱导的细胞凋亡，从而调节人肝癌细胞中Bcl-2:Bax的分子比例。
