AMPK-independent induction of autophagy by cytosolic Ca2+ increase

作者：Grotemeier Antje; Alers Sebastian; Pfisterer Simon G; Paasch Florian; Daubrawa Merle; Dieterle Alexandra; Viollet Benoit; Wesselborg Sebastian; Proikas Cezanne Tassula^*; Stork Bjoern

来源：Cellular Signalling, 2010, 22(6): 914-925.

DOI：10.1016/j.cellsig.2010.01.015

• Autophagy
• LC3
• Mammalian target of rapamycin
• Phosphoinositide-3 phosphate
• Thapsigargin
• WIPI-1

Autophagy is a eukaryotic lysosomal bulk degradation system initiated by cytosolic cargo sequestration in autophagosomes. The Ser/Thr kinase mTOR has been shown to constitute a central role in controlling the initiation of autophagy by integrating multiple nutrient-dependent signaling pathways that crucially involves the activity of PI3K class III to generate the phosphoinositide PI(3)P. Recent reports demonstrate that the increase in cytosolic Ca2+ can induce autophagy by inhibition of mTOR via the CaMKK-alpha/beta-mediated activation of AMPK. Here we demonstrate that Ca2+ signaling can additionally induce autophagy independently of the Ca2+-mediated activation of AMPK. First, by LC3-II protein monitoring in the absence or presence of lysosomal inhibitors we confirm that the elevation of cytosolic Ca2+ induces autophagosome generation and does not merely block autophagosome degradation. Further, we demonstrate that Ca2+-chelation strongly inhibits autophagy in human, mouse and chicken cells. Strikingly, we found that the PI(3)P-binding protein WIPI-1 (Atg18) responds to the increase of cytosolic Ca2+ by localizing to autophagosomal membranes (WIPI-1 puncta) and that Ca2+-chelation inhibits WIPI-1 puncta formation, although PI(3)P-generation is not generally affected by these Ca2+ flux modifications. Importantly, using AMPK-alpha 1(-/-)alpha 2(-/-) MEFs we show that thapsigargin application triggers autophagy in the absence of AMPK and does not involve complete mTOR inhibition, as detected by p7056K phosphorylation. In addition, STO-609-mediated CaMKK-alpha/beta inhibition decreased the level of thapsigargin-induced autophagy only in AMPK-positive cells. We suggest that apart from reported AMPK-dependent regulation of autophagic degradation, an AMPK-independent pathway triggers Ca2+-mediated autophagy, involving the PI(3)P-effector protein WIPI-1 and LC3.

• 出版日期2010-6