RIP3/MLKL 通过激活 4EBP1-eIF4E 通路来调节细胞坏死。
RIP3/MLKL regulates necroptosis via activating 4EBP1-eIF4E pathway.
机构信息
Center for Medical Research, Second Xiangya Hospital, Central South University, Changsha 410011.
Department of Gerontology, Second Xiangya Hospital, Central South University, Changsha 410011, China.
出版信息
Zhong Nan Da Xue Xue Bao Yi Xue Ban. 2023 Jul 28;48(7):979-985. doi: 10.11817/j.issn.1672-7347.2023.230153.
OBJECTIVES
Necroptosis is a cell death type mediated by receptor interacting protein 3 (RIP3)/mixed lineage kinase domain-like protein (MLKL). It has been reported that mammalian target of rapamycin plays a regulatory role in necroptosis. Eukaryotic translation initiation factor 4E-binding protein 1 (4EBP1)-eukaryotic initiation factor 4E (eIF4E) pathway is a key down streamer of mammalian target of rapamycin. However, whether 4EBP1-eIF4E pathway is involved in necroptosis is still unknown. This study aims to investigate the changes of 4EBP1-eIF4E pathway in necroptosis.
METHODS
TNF-α/SM-164/Z-VAD-FMK (TSZ), a necroptosis inducer, was used to induce necroptosis in murine fibroblastoid cell line L929. Cell necrosis was observed under an optical microscope. Then, TSZ was added to L929 cells with and gene knockout. Propidium iodide (PI) staining was used to observe cell necrosis. Real-time fluorescence quantitative PCR and Western blotting were used to determine the mRNA and protein expression of and , respectively.
RESULTS
After treating L929 cells with TSZ, the number of necrotic cells was increased, the mRNA and protein expression levels of were significantly downregulated, and the ratio of phosphorylated 4EBP1 (p-4EBP1) to 4EBP1 was increased (<0.05 or <0.01); the mRNA expression level of was significantly upregulated, and the ratio of phosphorylated eIF4E (p-eIF4E) to eIF4E was increased (both <0.01). After knocking out and in L929 cells, PI positive necrotic cells were significantly reduced, the mRNA and protein expression levels of were significantly upregulated, and the ratio of p-4EBP1 to 4EBP1 was decreased (<0.05 or <0.01); the mRNA expression level of was significantly downregulated, and the ratio of p-eIF4E to eIF4E was decreased (both <0.01).
CONCLUSIONS
4EBP1-eIF4E pathway is activated in the RIP3/MLKL mediated-necroptosis.
目的
坏死是一种由受体相互作用蛋白 3（RIP3）/混合谱系激酶结构域样蛋白（MLKL）介导的细胞死亡类型。据报道，哺乳动物雷帕霉素靶蛋白在坏死中起调节作用。真核翻译起始因子 4E 结合蛋白 1（4EBP1）-真核起始因子 4E（eIF4E）途径是哺乳动物雷帕霉素靶蛋白的关键下游通路。然而，4EBP1-eIF4E 途径是否参与坏死仍不清楚。本研究旨在探讨坏死中 4EBP1-eIF4E 途径的变化。
方法
使用 TNF-α/SM-164/Z-VAD-FMK（TSZ）诱导鼠成纤维样细胞系 L929 发生坏死。在光学显微镜下观察细胞坏死。然后，用 TSZ 处理 L929 细胞的 和 基因敲除细胞。用碘化丙啶（PI）染色观察细胞坏死。实时荧光定量 PCR 和 Western blot 分别用于检测 和 的 mRNA 和蛋白表达。
结果
用 TSZ 处理 L929 细胞后，坏死细胞数量增加， mRNA 和蛋白表达水平明显下调，磷酸化 4EBP1（p-4EBP1）与 4EBP1 的比值增加（<0.05 或 <0.01）； mRNA 表达水平明显上调，磷酸化 eIF4E（p-eIF4E）与 eIF4E 的比值增加（均<0.01）。在 L929 细胞中敲除 和 后，PI 阳性坏死细胞明显减少， mRNA 和蛋白表达水平明显上调，p-4EBP1 与 4EBP1 的比值降低（<0.05 或 <0.01）； mRNA 表达水平明显下调，p-eIF4E 与 eIF4E 的比值降低（均<0.01）。
结论
RIP3/MLKL 介导的坏死中激活了 4EBP1-eIF4E 通路。
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