Institute of Hansen's Disease, College of Medicine, The Catholic University of Korea, Seocho-gu, Seoul 137-701, Korea.
Free Radic Res. 2011 Apr;45(4):389-99. doi: 10.3109/10715762.2010.535530. Epub 2010 Nov 10.
A novel mechanism for H₂O₂-induced autophagic cell death in GSH-depleted RAW 264.7 cells, a murine macrophage cell line, is proposed. Under GSH-depleted conditions, H₂O₂-induced autophagic cell, characterized by an increased LC3-II/I ratio, a decreased level of p62 and the formation of autophagic vacuoles, was inhibited by bafilomycin A1 and by Atg5 siRNA transfection, whereas the cell death was not inhibited by zVAD-fmk, by PI3K inhibitors or by Beclin 1 siRNA transfection. In addition, H₂O₂ treatment reduced the activity of mTOR and promoted the ubiquitination and degradation of Rheb, a key upstream activator of mTOR. Furthermore, proteasome inhibition with MG132 restored the expression of Rheb and increased mTOR activity, resulting in an increased viability of H₂O₂-treated cells. Collectively, these findings demonstrate that H₂O₂ induces Beclin 1-independent autophagic cell death by suppressing the mTOR pathway via promoting the ubiquitination and degradation of Rheb in GSH-depleted RAW 264.7 cells.
提出了一种在 GSH 耗竭的 RAW 264.7 细胞（一种鼠巨噬细胞系）中 H₂O₂诱导自噬性细胞死亡的新机制。在 GSH 耗竭的条件下，H₂O₂诱导的自噬细胞，其特征是 LC3-II/I 比值增加、p62 水平降低和自噬小体的形成，被巴弗洛霉素 A1 和 Atg5 siRNA 转染抑制，而细胞死亡未被 zVAD-fmk、PI3K 抑制剂或 Beclin 1 siRNA 转染抑制。此外，H₂O₂处理降低了 mTOR 的活性，并促进了 Rheb 的泛素化和降解，Rheb 是 mTOR 的关键上游激活剂。此外，用 MG132 抑制蛋白酶体恢复了 Rheb 的表达并增加了 mTOR 活性，导致 H₂O₂处理的细胞活力增加。总之，这些发现表明，H₂O₂通过在 GSH 耗竭的 RAW 264.7 细胞中促进 Rheb 的泛素化和降解来抑制 mTOR 通路，从而诱导 Beclin 1 非依赖性自噬性细胞死亡。
