抑制聚ADP-核糖聚合酶-1活性降低HEK293/tau441细胞tau蛋白的磷酸化

Suppressing poly(ADP-ribose)polymerase-1 inhibits tau phosphorylation in HEK293/tau441 cells

为了研究聚ADP-核糖聚合酶-1[poly(ADP-ribose)polymerase-1,PARP-1]对微管相关蛋白tau磷酸化水平的影响,本实验分别用不同剂量(0.5,1,2,4mmol/L)的PARP-1的抑制剂3-氨基苯甲酰胺(3-aminobenzamide,3-AB)处理稳定表达tau441蛋白的HEK293细胞。24h后观察细胞的形态学变化,并用免疫印迹的方法检测PARP-1的聚ADP-核糖化修饰变化、微管相关蛋白tau的磷酸化水平和糖原合酶激酶-3(glycogen synthase kinase3,GSK-3)的活性改变情况。结果显示:(1)不同浓度的3-AB处理HEK293/tau441细胞后,细胞形态未发生显著的变化;(2)PARP-1的活性下降引起tau蛋白Ser195/198/199/202位点的非磷酸化增强;(3)PARP-1的活性抑制使tau蛋白Thr231位点的磷酸化下降,同时细胞内的GSK-3的活性也下降。结果提示,PARP-1的活性抑制可能通过下调GSK-3的活性,降低tau蛋白磷酸化水平。

The study aimed to investigate the effect of inhibition of poly（ADP-ribose） polymerase-1 （PARP-1） activity on tau phosphorylation in HEK293/tau441 cells and its mechanism. HEK293/tau441 cells were treated with 3-aminobenzamide （3-AB）, a PARP- 1 inhibitor, at different doses （0.5, 1, 2, 4 mmol/L）. After 24 h, the cell morphology was observed under phase contrast microscope, tau phosphorylation level in different sites （tau-1, tau-5, Thr231） and the activity of glycogen synthase kinase 3 （GSK-3） were detected by Western blotting. The results showed： （1） 3-AB at different doses failed to change the morphology of cells; （2） The 3-ABinduced decrease in activity of PARP-I resulted in increase of unphosphorylation level in tau-l（Ser195/198/199/202） sites; （3） The phosphorylation of tau was decreased in Thr231 site, while the total tau was slightly changed after 3-AB treatment; （4） With the increased phosphorylation of GSK-3 at Ser9 site, the activity of GSK-3 was decreased after 3-AB treatment. The results suggest that the inhibition of PARP-1 by 3-AB could decrease tau phosphorylation in HEK293/tau441 cells probably through decreasing GSK-3 activity.