ATR activation by Cr-DNA damage is a major survival response establishing late S and G2 checkpoints after Cr(VI) exposure.

Inhalation exposure to hexavalent chromium is known to cause lung cancer and other pulmonary toxicity. Cellular metabolism of chromium(VI) entering cells as chromate anion produces different amounts of reactive Cr(V) intermediates and finally yields Cr(III). Direct reduction of Cr(VI) by ascorbate (Asc), the dominant metabolic reaction in vivo but not in standard cell cultures, skips production of Cr(V) but still permits extensive formation of Cr-DNA damage. To understand the importance of different forms of biological injury in Cr(VI) toxicity, we examined activation of several protein- and DNA damage-sensitive stress responses in human lung cells under Asc-restored conditions. We found that Asc-restored cells suppressed upregulation of oxidant-sensitive stress systems by Cr(VI) but showed a strong activation of the apical DNA damage-responsive kinase ATR. ATR signaling was triggered in late S phase and persisted upon entry of cells into G2 phase. Inhibition of ATR prevented the establishment of late-S and G2 cell cycle checkpoints and did not lead to a compensatory activation of a related kinase ATM. Inactivation of ATR also strongly impaired viability of Cr(VI)-treated lung cells including stem-like cells and revealed a significant formation of toxic Cr-DNA damage at low Cr(VI) doses. Our findings identified a major Cr(VI) resistance mechanism involving sensing of Cr-DNA damage by ATR in late S phase and a subsequent establishment of protective cell cycle checkpoints.