Division of Biochemistry, College of Pharmacy, Chung-Ang University, 221 Huksuk-Dong, Dongjak-Gu, Seoul, Korea.
J Toxicol Environ Health A. 2010;73(21-22):1465-76. doi: 10.1080/15287394.2010.511553.
Ceramide induces cell cycle arrest and apoptotic cell death associated with increased levels of p27(kip1). The aim of this study was to examine the effects of ceramide on p27(kip1) protein levels as a measure of cell cycle arrest and apoptosis. Results showed that ceramide increased p27(kip1) protein levels through activation of protein phosphatase 2A (PP2A) in PC-3 prostate cancer cells. Treatment of cells with the PP2A inhibitor okadaic acid or with PP2A-Cα siRNA inhibited ceramide-induced enhanced p27(kip1) protein expression and Akt dephosphorylation, and prevented Skp2 downregulation. Overexpression of constitutively active Akt attenuated ceramide-induced Skp2 downregulation and p27(kip1) upregulation. In addition, ceramide stimulated binding of the PP2A catalytic subunit PP2A-Cαβ to Akt as assessed by immunoprecipitation experiments, indicating that PP2A is involved in the induction of p27(kip1) via inhibition of Akt pathway. Finally, whether PP2A can regulate p27(kip1) expression independently of Akt pathway was determined. Knockdown of PP2A-Cα with siRNA reduced p27(kip1) levels in the presence of Akt inhibitor. These data reveal that PP2A is a regulator of ceramide-induced p27(kip1) expression via Akt-dependent and Akt-independent pathways.
神经酰胺通过激活蛋白磷酸酶 2A（PP2A）诱导细胞周期停滞和与 p27（kip1）水平升高相关的细胞凋亡。本研究旨在研究神经酰胺对 p27（kip1）蛋白水平的影响，作为细胞周期停滞和凋亡的衡量标准。结果表明，神经酰胺通过激活蛋白磷酸酶 2A（PP2A）在 PC-3 前列腺癌细胞中增加 p27（kip1）蛋白水平。用 PP2A 抑制剂 okadaic 酸或 PP2A-Cα siRNA 处理细胞，抑制了神经酰胺诱导的增强的 p27（kip1）蛋白表达和 Akt 去磷酸化，并阻止了 Skp2 下调。组成型激活 Akt 的过表达减弱了神经酰胺诱导的 Skp2 下调和 p27（kip1）上调。此外，通过免疫沉淀实验评估，神经酰胺刺激了 PP2A 催化亚基 PP2A-Cαβ与 Akt 的结合，表明 PP2A 通过抑制 Akt 途径参与诱导 p27（kip1）。最后，确定了 PP2A 是否可以独立于 Akt 途径调节 p27（kip1）的表达。用 siRNA 敲低 PP2A-Cα 可降低 Akt 抑制剂存在时的 p27（kip1）水平。这些数据表明，PP2A 通过 Akt 依赖性和非依赖性途径调节神经酰胺诱导的 p27（kip1）表达。
