Modulation of intracellular calcium homeostasis blocks autophagosome formation

Nikolai Engedal, Maria L Torgersen, Ingrid J Guldvik, Stefan J Barfeld, Daniela Bakula, Frank Sætre, Linda K Hagen, John B Patterson, Tassula Proikas-Cezanne, Per O Seglen, Anne Simonsen, Ian G Mills

Research output: Contribution to journalArticlepeer-review

57 Citations (Scopus)

Abstract

Cellular stress responses often involve elevation of cytosolic calcium levels, and this has been suggested to stimulate autophagy. Here, however, we demonstrated that agents that alter intracellular calcium ion homeostasis and induce ER stress-the calcium ionophore A23187 and the sarco/endoplasmic reticulum Ca (2+)-ATPase inhibitor thapsigargin (TG)-potently inhibit autophagy. This anti-autophagic effect occurred under both nutrient-rich and amino acid starvation conditions, and was reflected by a strong reduction in autophagic degradation of long-lived proteins. Furthermore, we found that the calcium-modulating agents inhibited autophagosome biogenesis at a step after the acquisition of WIPI1, but prior to the closure of the autophagosome. The latter was evident from the virtually complete inability of A23187- or TG-treated cells to sequester cytosolic lactate dehydrogenase. Moreover, we observed a decrease in both the number and size of starvation-induced EGFP-LC3 puncta as well as reduced numbers of mRFP-LC3 puncta in a tandem fluorescent mRFP-EGFP-LC3 cell line. The anti-autophagic effect of A23187 and TG was independent of ER stress, as chemical or siRNA-mediated inhibition of the unfolded protein response did not alter the ability of the calcium modulators to block autophagy. Finally, and remarkably, we found that the anti-autophagic activity of the calcium modulators did not require sustained or bulk changes in cytosolic calcium levels. In conclusion, we propose that local perturbations in intracellular calcium levels can exert inhibitory effects on autophagy at the stage of autophagosome expansion and closure.

Original languageEnglish
Pages (from-to)1475-90
Number of pages16
JournalAutophagy
Volume9
Issue number10
DOIs
Publication statusPublished - Oct 2013

Keywords

  • Autophagy
  • Calcium
  • Cell Communication
  • Cell Line
  • Cytosol
  • Endoplasmic Reticulum
  • Homeostasis
  • Humans
  • Intracellular Space
  • Signal Transduction
  • Thapsigargin

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