Kim Hye-Gyo, Ro Myeong-Han, Lee Michael
Division of Life Sciences, College of Life Sciences and Bioengineering, Incheon National University, 119 Academy-Ro, Yeonsu-Gu, Incheon, 22012 Republic of Korea.
Institute for New Drug Development, Incheon National University, Incheon, 22012 Republic of Korea.
Toxicol Res. 2023 Jun 5;39(4):637-647. doi: 10.1007/s43188-023-00191-3. eCollection 2023 Oct.
Autophagy play contradictory roles in cellular transformation. We previously found that the knockout (KO) of autophagy-related 5 (Atg5), which is essential for autophagy, leads to the malignant transformation of NIH 3T3 cells. In this study, we explored the mechanism by which autophagy contributes to this malignant transformation using two transformed cell lines, Atg5 KO and Ras-NIH 3T3. Monomeric red fluorescent protein-green fluorescent protein-light chain 3 reporter and Cyto-ID staining revealed that Ras-NIH 3T3 cells exhibited higher basal autophagy activity than NIH 3T3 cells. Additionally, transformed cells, regardless of their Atg5 KO status, were more sensitive to autophagy inhibitors (SBI-0206965, chloroquine, and obatoclax) than the untransformed NIH 3T3 cells, suggesting that the transformed cells are more autophagy-dependent than the normal cells. Loss of Atg5 improved the cell viability and mobility, especially in Ras-NIH 3T3 cells. Furthermore, we discovered that autophagy was alternatively induced in a Rab9-dependent manner in Ras-NIH 3T3 and NIH 3T3/Atg5 KO cells. In particular, Atg5 KO cells showed reduced mTOR-mediated phosphorylation of Akt (pAkt S473), indicating the mTOR-independent occurrence of alternative autophagy in Atg5 KO cells. Therefore, our study provides evidence that alternative autophagy may contribute to tumorigenesis in cells with an impaired Atg5-dependent autophagy pathway.
The online version contains supplementary material available at 10.1007/s43188-023-00191-3.
自噬在细胞转化中发挥着矛盾的作用。我们之前发现，对自噬至关重要的自噬相关5（Atg5）基因敲除（KO）会导致NIH 3T3细胞发生恶性转化。在本研究中，我们使用两种转化细胞系Atg5 KO和Ras-NIH 3T3，探究了自噬促成这种恶性转化的机制。单体红色荧光蛋白-绿色荧光蛋白-轻链3报告基因和Cyto-ID染色显示，Ras-NIH 3T3细胞的基础自噬活性高于NIH 3T3细胞。此外，无论Atg5 KO状态如何，转化细胞比未转化的NIH 3T3细胞对自噬抑制剂（SBI-0206965、氯喹和 obatoclax）更敏感，这表明转化细胞比正常细胞更依赖自噬。Atg5的缺失提高了细胞活力和迁移能力，尤其是在Ras-NIH 3T3细胞中。此外，我们发现自噬在Ras-NIH 3T3和NIH 3T3/Atg5 KO细胞中以Rab9依赖的方式被选择性诱导。特别是，Atg5 KO细胞显示出mTOR介导的Akt磷酸化（pAkt S473）减少，表明Atg5 KO细胞中存在不依赖mTOR的选择性自噬。因此，我们的研究提供了证据，表明选择性自噬可能在Atg5依赖的自噬途径受损的细胞中促进肿瘤发生。
在线版本包含可在10.1007/s43188-023-00191-3获取的补充材料。
