Identification of Kinases and Phosphatases That Regulate ATG4B Activity by siRNA and Small Molecule Screening in Cells.
Identification of Kinases and Phosphatases That Regulate ATG4B Activity by siRNA and Small Molecule Screening in Cells.
复制标题
鉴定siRNA和小分子筛选细胞中ATG4B活性的激酶和磷酸酶。
DOI：
10.3389/fcell.2018.00148
复制
发表时间：
2018
影响因子：
5.5
通讯作者：
Ketteler R
中科院分区：
文献类型：
--
作者：
Pengo N;Prak K;Costa JR;Luft C;Agrotis A;Freeman J;Gewinner CA;Chan AWE;Selwood DL;Kriston-Vizi J;Ketteler R
Autophagy protease ATG4B is a key regulator of the LC3/GABARAP conjugation system required for autophagosome formation, maturation and closure. Members of the ATG4 and the LC3/GABARAP family have been implicated in various diseases including cancer, and targeting the ATG4B protease has been suggested as a potential therapeutic anti-cancer strategy. Recently, it has been demonstrated that ATG4B is regulated by multiple post-translational modifications, including phosphorylation and de-phosphorylation. In order to identify regulators of ATG4B activity, we optimized a cell-based luciferase assay based on ATG4B-dependent release of Gaussia luciferase. We applied this assay in a proof-of-concept small molecule compound screen and identified activating compounds that increase cellular ATG4B activity. Next, we performed a high-throughput screen to identify kinases and phosphatases that regulate cellular ATG4B activity using siRNA mediated knockdown and cDNA overexpression. Of these, we provide preliminary evidence that the kinase AKT2 enhances ATG4B activity in cells. We provide all raw and processed data from the screens as a resource for further analysis. Overall, our findings provide novel insights into the regulation of ATG4B and highlight the importance of post-translational modifications of ATG4B.
