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 (mitoO(2)(-& BULL;)) via a Ca2+-dependent mechanism, in which an initial stimulation of the inositol 1, 4, 5trisphosphate 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 1 alpha (ERO1 alpha) levels in proliferating U937 cells. EN460, an inhibitor of ERO1 alpha, recapitulated all the effects associated with RyR inhibition or downregulation, including prevention of RyR-induced Ca2+ accumulation in mitochondria and the resulting O-2(-& BULL;) formation. Quantitatively similar results were obtained in inhibitor studies performed in terminally differentiated wild type C2C12 cells. Moreover, ERO1 alpha knockout C2C12 myotubes responded to arsenite as their wild type counterpart supplemented with EN460. As a final note, arsenite enhanced the expression of ERO1 alpha 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 alpha in the ER, by which IP3R-dependent Ca2+ induces ERO1 alpha and ERO1 alpha promotes Ca2+ release via RyR, thereby amplifying the initial Ca2+ load and causing the mitochondrial accumulation of the cation, critical for mitoO(2)(-& BULL;) formation.