Crosstalk between ERO1α and ryanodine receptor in arsenite-dependent mitochondrial ROS formation

Arsenite, a well-established human carcinogen and toxic compound, promotes the formation of mitochondrial superoxide (mitoO₂^-) via a Ca²⁺-dependent mechanism, in which an initial stimulation of the inositol 1, 4, 5-trisphosphate receptor (IP₃R) is followed by the activation of the ryanodine receptor (RyR), critical for providing Ca²⁺ to the mitochondria. We now report that, under the same conditions, arsenite triggers endoplasmic reticulum (ER) stress and a threefold increase in ER oxidoreductin 1α (ERO1 α) levels in proliferating U937 cells. EN460, an inhibitor of ERO1 α, recapitulated all the effects associated with RyR inhibition or downregulation, including prevention of RyR-induced Ca²⁺ accumulation in mitochondria and the resulting O₂^-. formation. Quantitatively similar results were obtained in inhibitor studies performed in terminally differentiated wild type C2C12 cells. Moreover, ERO1 α knockout C2C12 myotubes responded to arsenite as their wild type counterpart supplemented with EN460. As a final note, arsenite enhanced the expression of ERO1 α via a mechanism mediated by Ca²⁺ release from both the IP₃R and RyR. We therefore conclude that arsenite activates a positive feedback amplification cycle between Ca²⁺ levels and ERO1 α in the ER, by which IP₃R-dependent Ca²⁺ induces ERO1 α and ERO1 α promotes Ca²⁺ release via RyR, thereby amplifying the initial Ca²⁺ load and causing the mitochondrial accumulation of the cation, critical for mitoO₂^- formation.