Targeted disruption of GAK stagnates autophagic flux by disturbing lysosomal dynamics
Targeted disruption of GAK stagnates autophagic flux by disturbing lysosomal dynamics
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DOI：
10.3892/ijmm.2021.5028
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发表时间：
2021-10-01
影响因子：
5.4
通讯作者：
Miyazawa, Keisuke
中科院分区：
文献类型：
--
作者：
Miyazaki, Masaya;Hiramoto, Masaki;Miyazawa, Keisuke
自噬-溶酶体系统允许细胞通过调节细胞组分的降解和再循环来适应环境变化，并通过去除聚集的蛋白质和有缺陷的细胞器来维持稳态。细胞周期蛋白G相关激酶（GAK）参与调节网格蛋白依赖的内吞作用和细胞周期进程。此外，在GAK基因座的单核苷酸多态性已被报道为帕金森病的危险因素。然而，GAK在自噬-溶酶体系统中的作用还不完全清楚，因此本研究旨在阐明这一点。在本研究中，在遗传破坏或化学抑制GAK的情况下，分析自噬通量和观察自噬体和自溶酶体的形态学变化表明，GAK通过肌动球蛋白调节来控制溶酶体动力学，导致自噬的稳定进行。GAK基因敲除（KO）可抑制A549细胞自噬体-溶酶体融合和自噬溶酶体重组，导致自噬体和自噬溶酶体在长期饥饿条件下大量积累。伴随着这些现象的自噬通量的停滞也观察到与GAK抑制剂的加入。此外，Rho相关蛋白激酶（ROCK）抑制剂或ROCK 1敲低的加入减轻了GAK KO介导的作用。结果表明，GAK在自噬过程中通过维持溶酶体的稳态来控制溶酶体动力学的重要作用。
The autophagy-lysosome system allows cells to adapt to environmental changes by regulating the degradation and recycling of cellular components, and to maintain homeostasis by removing aggregated proteins and defective organelles. Cyclin G-associated kinase (GAK) is involved in the regulation of clathrin-dependent endocytosis and cell cycle progression. In addition, a single nucleotide polymorphism at the GAK locus has been reported as a risk factor for Parkinson's disease. However, the roles of GAK in the autophagy-lysosome system are not completely understood, thus the present study aimed to clarify this. In the present study, under genetic disruption or chemical inhibition of GAK, analyzing autophagic flux and observing morphological changes of autophagosomes and autolysosomes revealed that GAK controlled lysosomal dynamics via actomyosin regulation, resulting in a steady progression of autophagy. GAK knockout (KO) in A549 cells impaired autophagosome-lysosome fusion and autophagic lysosome reformation, which resulted in the accumulation of enlarged autophagosomes and autolysosomes during prolonged starvation. The stagnation of autophagic flux accompanied by these phenomena was also observed with the addition of a GAK inhibitor. Furthermore, the addition of Rho-associated protein kinase (ROCK) inhibitor or ROCK1 knockdown mitigated GAK KO-mediated effects. The results suggested a vital role of GAK in controlling lysosomal dynamics via maintaining lysosomal homeostasis during autophagy.