Resveratrol-induced autophagy is dependent on IP3Rs and on cytosolic Ca2 +☆

Abstract Previous work revealed that intracellular Ca 2 + signals and the inositol 1,4,5-trisphosphate (IP 3 ) receptors (IP 3 R) are essential to increase autophagic flux in response to mTOR inhibition, induced by either nutrient starvation or rapamycin treatment. Here, we investigated whether autophagy induced by resveratrol, a polyphenolic phytochemical reported to trigger autophagy in a non-canonical way, also requires IP 3 Rs and Ca 2 + signaling. Resveratrol augmented autophagic flux in a time-dependent manner in HeLa cells. Importantly, autophagy induced by resveratrol (80 μM, 2 h) was completely abolished in the presence of 10 μM BAPTA-AM, an intracellular Ca 2 + -chelating agent. To elucidate the IP 3 R's role in this process, we employed the recently established HEK 3KO cells lacking all three IP 3 R isoforms. In contrast to the HEK293 wt cells and to HEK 3KO cells re-expressing IP 3 R1, autophagic responses in HEK 3KO cells exposed to resveratrol were severely impaired. These altered autophagic responses could not be attributed to alterations in the mTOR/p70S6K pathway, since resveratrol-induced inhibition of S6 phosphorylation was not abrogated by chelating cytosolic Ca 2 + or by knocking out IP 3 Rs. Finally, we investigated whether resveratrol by itself induced Ca 2 + release. In permeabilized HeLa cells, resveratrol neither affected the sarco- and endoplasmic reticulum Ca 2 + ATPase (SERCA) activity nor the IP 3 -induced Ca 2 + release nor the basal Ca 2 + leak from the ER. Also, prolonged (4 h) treatment with 100 μM resveratrol did not affect subsequent IP 3 -induced Ca 2 + release. However, in intact HeLa cells, although resveratrol did not elicit cytosolic Ca 2 + signals by itself, it acutely decreased the ER Ca 2 + -store content irrespective of the presence or absence of IP 3 Rs, leading to a dampened agonist-induced Ca 2 + signaling. In conclusion, these results reveal that IP 3 Rs and cytosolic Ca 2 + signaling are fundamentally important for driving autophagic flux, not only in response to mTOR inhibition but also in response to non-canonical autophagy inducers like resveratrol. This article is part of a Special Issue entitled: ECS Meeting edited by Claus Heizmann, Joachim Krebs and Jacques Haiech.