ATX-LPA-Dependent Nuclear Translocation of Endonuclease G in Respiratory Epithelial Cells: A New Mode Action for DNA Damage Induced by Crystalline Silica Particles

SIMPLE SUMMARY: Crystalline silica particles (CSi) are carcinogenic; however, their genotoxicity is not well characterized. Indirect evidence suggests that retained particles in peripheral bronchioles activate macrophages, causing chronic ROS production. Using human bronchial epithelia in monocultures, we have recently shown that CSi activate ATX at the plasma membrane and that a complex signaling pathway leads to double strand breaks (DSBs). The DSBs were seen within minutes, also in mice inhaling CSi. A remaining question is how DSBs are induced. Using the same models as previously, we now show that ATX signaling leads to endonuclease G translocation from the mitochondria to the nucleus. It was synchronized with DSB formation, and inhibitors prevented both the translocation and the DSBs, supporting a causal role for endonuclease G. The CSi-induced micronuclei formation indicated genomic instability. Additionally, the endonuclease G mediated genotoxicity, which thus affects the respiratory epithelium directly at very low particle doses, might explain the carcinogenic effects of CSi, perhaps even better than ROS-producing macrophages. ABSTRACT: Crystalline silica particles (CSi) are an established human carcinogen, but it is not clear how these particles cause necessary mutations. A well-established scenario includes inflammation caused by retained particles in the bronchioles, activated macrophages, and reactive oxygen species (ROS) that cause DNA damage. In previous studies, we showed that CSi in contact with the plasma membrane of human bronchial epithelium induced double strand breaks within minutes. A signaling pathway implicating the ATX-LPA axis, Rac1, NLRP3, and mitochondrial depolarization upstream of DSB formation was delineated. In this paper, we provide in vitro and in vivo evidence that this signaling pathway triggers endonuclease G (EndoG) translocation from the mitochondria to the nucleus. The DNA damage is documented as γH2AX and p53BP1 nuclear foci, strand breaks in the Comet assay, and as micronuclei. In addition, the DNA damage is induced by low doses of CSi that do not induce apoptosis. By inhibiting the ATX-LPA axis or by EndoG knockdown, we prevent EndoG translocation and DSB formation. Our data indicate that CSi in low doses induces DSBs by sub-apoptotic activation of EndoG, adding CSi to a list of carcinogens that may induce mutations via sub-apoptotic and “minority MOMP” effects. This is the first report linking the ATX-LPA axis to this type of carcinogenic effect.