Ca2+ mobilization-dependent reduction of the endoplasmic reticulum lumen is due to influx of cytosolic glutathione
Beáta Lizák, Julia Birk, Melinda Zana, Gergely Kosztyi, Denise V. Kratschmar, Alex Odermatt, Richard Zimmermann, Miklós Geiszt, Christian Appenzeller‐Herzog, Gábor Bánhegyi
Abstract
Abstract Background The lumen of the endoplasmic reticulum (ER) acts as a cellular Ca 2+ store and a site for oxidative protein folding, which is controlled by the reduced glutathione (GSH) and glutathione-disulfide (GSSG) redox pair. Although depletion of luminal Ca 2+ from the ER provokes a rapid and reversible shift towards a more reducing poise in the ER, the underlying molecular basis remains unclear. Results We found that Ca 2+ mobilization-dependent ER luminal reduction was sensitive to inhibition of GSH synthesis or dilution of cytosolic GSH by selective permeabilization of the plasma membrane. A glutathione-centered mechanism was further indicated by increased ER luminal glutathione levels in response to Ca 2+ efflux. Inducible reduction of the ER lumen by GSH flux was independent of the Ca 2+ -binding chaperone calreticulin, which has previously been implicated in this process. However, opening the translocon channel by puromycin or addition of cyclosporine A mimicked the GSH-related effect of Ca 2+ mobilization. While the action of puromycin was ascribable to Ca 2+ leakage from the ER, the mechanism of cyclosporine A-induced GSH flux was independent of calcineurin and cyclophilins A and B and remained unclear. Conclusions Our data strongly suggest that ER influx of cytosolic GSH, rather than inhibition of local oxidoreductases, is responsible for the reductive shift upon Ca 2+ mobilization. We postulate the existence of a Ca 2+ - and cyclosporine A-sensitive GSH transporter in the ER membrane. These findings have important implications for ER redox homeostasis under normal physiology and ER stress.