Dynactin调控α-微管蛋白乙酰化和微管修饰的机制研究

The mechanism of Dynactin in mediating alpha-tubulin acetylation and tubulin modification

目的探讨动力蛋白激活蛋白Dynactin对α-微管蛋白乙酰化水平和微管修饰的调控作用。方法利用蛋白印迹方法检测Dynactin亚基p150~(Glued)N端缺失(KO)和对照(Ctrl)小鼠原代皮质神经元和原代成纤维细胞中α-微管蛋白乙酰化水平和总蛋白水平。利用免疫荧光染色方法观察Ctrl和KO原代成纤维细胞中微管乙酰化百分比的变化情况。结果p150~(Glued)N端缺失不影响原代皮质神经元中Dynactin及乙酰化α-微管蛋白水平。与Ctrl小鼠原代成纤维细胞比,KO原代成纤维细胞中Dynactin亚基蛋白水平降低,且乙酰化α-微管蛋白水平明显升高。此外在KO原代成纤维细胞中也观察到微管乙酰化的百分比显著增多。结论Dynactin功能缺失可显著上调乙酰化α-微管蛋白水平,增加细胞微管乙酰化的百分比。

Objective To investigate the effect of dynactin onα-tubulin acetylation and tubulin modification.Methods Western blot was applied to detect the levels of acetylatedα-tubulin in the primary cultured cortical neuron and fibroblasts of dynactin subunit p150~(Glued)depletion(KO)and control(Ctrl)mice.The percentage of acetylated tubulin in Ctrl and KO primary cultured fibroblasts was observed by using immunofluorescence staining.Results Dynactin p150~(Glued)depletion have no changes on the levels of Dynactin subunits and acetylatedα-tubulin.Compared with the Ctrl fibroblasts,the protein level of Dynactin subunits was diseased and the level of acetylatedα-tubulin was increased in the KO fibroblasts.Moreover,the percentage of acetylated tubulin was increased in the KO fibroblasts.Conclusion Loss of function of Dynactin resulted in the increase of acetylatedα-tubulin level and tubulin acetylation in the primary cultured fibroblasts.