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Inhibition of calcium release from InsP3Rs triggers autophagy

Chehab, Tala; Rietdorf, Katja; Parys, Jan; Bultynck, Geert and Bootman, Martin (2015). Inhibition of calcium release from InsP3Rs triggers autophagy. In: Cellular Signalling Abstracts, article no. PC241.

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Abstract

Macroautophagy (autophagy) is a cellular self-eating process that is important for cell survival. It involves degradation of misfolded proteins, dysfunctional organelles and pathogens through the action of lysosomal hydrolases. In eukaryotic cells, autophagy has a housekeeping role in cellular turnover, and is substantially increased in response to nutrient starvation. A growing body of evidence indicates that dysregulation of autophagy is linked to a number of disorders, such as Alzheimer’s disease, cancer and heart diseases. Intracellular calcium has been established as a regulator of basal and starvation-induced autophagy. The mechanisms by which calcium regulates autophagy are not fully clear. It has been suggested that calcium signals can have both pro-autophagic and anti-autophagic effects.

A convenient method for assessing cellular autophagy in living cells is to monitor the generation of autophagic vesicles (AVs) using fluorescent reporters. In this study, HeLa cells stably expressing GFP-tagged microtubule-associated protein 1A/1B-light chain 3 (GFP-LC3), or wild type HeLa cells stained with a Cyto-ID autophagy detection kit (Enzo Life Sciences), were used to quantitate the formation of AVs in response to various stimuli. We found that starvation of HeLa cells in nutrient-deficient media or treatment with rapamycin (1 µM; a mTOR inhibitor) enhanced autophagy in a time- and concentration-dependent manner (17 ± 1.3 vs. 28 ± 1.5 AVs for basal vs. 1 hour nutrient starvation; and 17 ± 1.3 vs. 27 ± 1.9 AVs for basal vs. rapamycin treatment). Preventing calcium release from inositol 1,4,5-trisphosphate receptors (InsP3Rs) using expression of an InsP3 5’- phosphatase enzyme (5’-Pase) or 2-aminoethoxydiphenyl borate (100 µM; 2-APB) also resulted in enhanced autophagy (17 ± 1.3 vs. 28 ± 3.9 AVs for basal vs. 5’-Pase expression). The level of autophagy evoked by inhibition of InsP3R-mediated calcium release was similar to that caused by nutrient starvation or rapamycin.

Importantly, the extent of autophagy induced by nutrient starvation, rapamycin or 5’-Pase expression was enhanced by bafilomyin (0.1 µM), which inhibits the lysosomal processing of autophagic vesicles. This finding confirms that all three treatments increased the autophagic flux in HeLa cells. Our data suggest that calcium release from InsP3Rs plays a role in controlling the induction of autophagy. Plausibly, the release of calcium from InsP3Rs is required to stimulate mitochondrial respiration and ATP production. To test this hypothesis, we inhibited mitochondrial respiration with oligomycin (0.5 µM) and found an increase in autophagy in both cases (17 ± 1.2 vs. 26 ± 1.5 AVs for basal vs. oligomycin treatment). The results of our study are consistent with the proposal that the flux of calcium from InsP3Rs to the mitochondrial matrix is necessary to stimulate ATP production and avoid induction of autophagy.

Item Type: Conference or Workshop Item
Copyright Holders: 2015 The Authors
Academic Unit/School: Faculty of Science, Technology, Engineering and Mathematics (STEM) > Life, Health and Chemical Sciences
Faculty of Science, Technology, Engineering and Mathematics (STEM)
Interdisciplinary Research Centre: Biomedical Research Network (BRN)
Health and Wellbeing PRA (Priority Research Area)
Related URLs:
Item ID: 44174
Depositing User: Katja Rietdorf
Date Deposited: 17 Sep 2015 09:05
Last Modified: 11 Sep 2017 15:32
URI: http://oro.open.ac.uk/id/eprint/44174
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