Radiosensitization of radioresistant cancer stem cells by inhibition of ATM

我们在此报道，ATM可以在暴露于过氧化氢时被激活，而不会诱导Hep G2细胞的核DDR，并且氧化的细胞可以进行亚细胞分离。第一个基于洗涤剂的分离实验表明，活性磷酸化的ATM位于第二个部分，其中也含有线粒体和过氧化物酶体。另一种涉及均质和差速离心的分离方法允许产生含有过氧化物酶体的轻膜组分，但不产生线粒体，表明轻膜组分仅含有微量的ATM。相比之下，以线粒体成分为主的重膜组分富集了ATM和活性ATM，说明ATM的氧化活化发生在线粒体而不是过氧化物酶体中。在Rho 0-Hep G2细胞中，缺乏线粒体DNA和功能性线粒体，ATM对过氧化氢没有反应。

We report herein that ATM can be activated when exposed to hydrogen peroxide without inducing nuclear DDR in Hep G2 cells, and the oxidized cells could be subjected to subcellular fractionation. The first detergent-based fractionation experiment revealed that active, phosphorylated ATM was located in the second fraction, which also contained both mitochondria and peroxisomes. An alternative fractionation method involving homogenization and differential centrifugation, which permits the light membrane fraction containing peroxisomes to be produced, but not mitochondria, revealed that the light membrane fraction contained only traces of ATM. In contrast, the heavy membrane fraction, which mainly contained mitochondrial components, was enriched in ATM and active ATM, suggesting that the oxidative activation of ATM occurs in mitochondria and not in peroxisomes. In Rho 0-Hep G2 cells, which lack mitochondrial DNA and functional mitochondria, ATM failed to respond to hydrogen peroxide.