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目的:通过建立过表达PC-1的前列腺癌LNCaP细胞系及敲低PC-1表达的C4-2细胞系,探究PC-1激活AKT信号通路的分子机制。方法:将PC-1基因及针对PC-1的siRNA序列,分别克隆至慢病毒表达载体pCDH-EF1-Myc-MCS-T2A-Puro及干扰载体pSIH1-H1-Puro,包装成慢病毒后分别感染前列腺癌LNCaP及C4-2细胞,通过Western印迹鉴定PC-1过表达及敲低效果,并检测PI3K/AKT/mTOR信号通路相关蛋白S6K、AKT的磷酸化水平。结果:PC-1过表达时,S6K磷酸化水平下降,而AKT的磷酸化水平上升。结论:PC-1可以通过抑制S6K激酶活性,解除其对AKT的负反馈抑制作用,从而激活AKT激酶的活性。
OBJECTIVE: To explore the molecular mechanism of PC-1 activation of AKT signaling by establishing prostate cancer LNCaP cell line overexpressing PC-1 and knockdown of PC-1 expressing C4-2 cell line. Methods: The PC-1 gene and the siRNA sequence targeting PC-1 were cloned into the lentiviral vector pCDH-EF1-Myc-MCS-T2A-Puro and the interference vector pSIH1-H1-Puro respectively. Prostate cancer LNCaP and C4-2 cells were used to detect the effect of PC-1 overexpression and knockdown by Western blotting. The phosphorylation of S6K and AKT were detected by Western blot. Results: When PC-1 was overexpressed, phosphorylation of S6K decreased and phosphorylation of AKT increased. Conclusion: PC-1 can activate AKT kinase by inhibiting S6K kinase activity and releasing its negative feedback inhibition on AKT.