Auxin-mediated rapid degradation of target proteins in hippocampal neurons
Auxin-mediated rapid degradation of target proteins in hippocampal neurons
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DOI：
10.1097/wnr.0000000000001299
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发表时间：
2019-09-04
期刊：
影响因子：
1.7
通讯作者：
Takeuchi, Haruki
中科院分区：
文献类型：
--
作者：
Nakano, Risako;Ihara, Naoki;Takeuchi, Haruki
蛋白质水平的遗传操作是一种很有前途的方法，以确定生物体中特定蛋白质的功能。先前的研究表明，生长素诱导的降解决定子策略提供了真菌和哺乳动物有丝分裂细胞中的各种蛋白质的快速和可逆的降解。在这项研究中，我们将这项技术应用于有丝分裂后的神经元，以解决生长素诱导的降解决定子系统是否可以应用于神经系统。利用腺相关病毒，我们同时引入增强型绿色荧光蛋白（EGFP）融合生长素诱导降解决定子标签和F-box家族蛋白，水稻（OsTIR 1），从小鼠海马神经元。在分离的海马神经元中，EGFP增强的绿色荧光蛋白荧光信号在加入植物激素生长素后迅速减弱。此外，生长素诱导的增强绿色荧光蛋白降解也观察到在海马急性脑片。总之，这些结果为神经科学家打开了一扇大门，以时间控制的方式通过生长素诱导的降解决定子系统操纵蛋白质表达水平。
Genetic manipulation of protein levels is a promising approach to identify the function of a specific protein in living organisms. Previous studies demonstrated that the auxin-inducible degron strategy provides rapid and reversible degradation of various proteins in fungi and mammalian mitotic cells. In this study, we employed this technology to postmitotic neurons to address whether the auxin-inducible degron system could be applied to the nervous system. Using adeno-associated viruses, we simultaneously introduced enhanced green fluorescent protein (EGFP) fused with an auxin-inducible degron tag and an F-box family protein, TIR1 from Oryza sativa (OsTIR1), into hippocampal neurons from mice. In dissociated hippocampal neurons, EGFP enhanced green fluorescent protein fluorescence signals rapidly decreased when adding a plant hormone, auxin. Furthermore, auxin-induced enhanced green fluorescent protein degradation was also observed in hippocampal acute slices. Taken together, these results open the door for neuroscientists to manipulate protein expression levels by the auxin-inducible degron system in a temporally controlled manner.