High Levels of ROS Impair Lysosomal Acidity and Autophagy Flux in Glucose-Deprived Fibroblasts by Activating ATM and Erk Pathways.

Seon Beom Song,Eun Seong Hwang

doi:10.3390/biom10050761

Abstract

Under glucose deprivation, cells heavily mobilize oxidative phosphorylation to maintain energy homeostasis. This leads to the generation of high levels of ATP, as well as reactive oxygen species (ROS), from mitochondria. In nutrient starvation, autophagy is activated, likely to facilitate resource recycling, but recent studies suggest that autophagy flux is inhibited in cells undergoing glucose deprivation. In this study, we analyzed the status of autophagic flux in glucose-deprived human fibroblasts. Although lysosomes increased in quantity due in part to an increase of biogenesis, a large population of them suffered low acidity in the glucose-deprived cells. Autophagosomes also accumulated due to poor autolysis in these cells. A treatment of antioxidants not only restored lysosomal acidity but also released the flux blockade. The inhibition of ataxia telangiectasia mutated (ATM) serine/threonine kinase, which is activated by ROS, also attenuated the impairment of lysosomal acidity and autophagic flux, suggesting an effect of ROS that might be mediated through ATM activation. In addition, the activity of extracellular signal-regulated kinase (Erk) increased upon glucose deprivation, but this was also compromised by a treatment of antioxidants. Furthermore, the Erk inhibitor treatment also alleviated the failure in lysosomal acidity and autophagic flux. These together indicate that, upon glucose deprivation, cells undergo a failure of autophagy flux through an impairment of lysosomal acidity and that a high-level ROS-induced activation of Erk and ATM is involved in this impairment.

Highlights

Autophagy is an important cellular salvage mechanism of recycling resources for anabolic building blocks, as well as energy
We examined the status of autophagic flux in human fibroblasts under glucose deprivation and confirmed these findings
In glucose-deprived cells, the levels of LC3 type II molecules (LC3-II) increased steadily over 72 h, and bafilomycin A1 (bafA1) treatment did not further elevate the levels at each time point

Summary

Introduction

Autophagy is an important cellular salvage mechanism of recycling resources for anabolic building blocks, as well as energy. Recent studies demonstrated that, in glucose-deprived cells, fatty acid β-oxidation and mitochondrial oxidative phosphorylation (OXPHOS) are increased [9,10,11,12,13,14,15] These changes would help cells maintain energy homeostasis in the absence of glucose input [11,16], but the activation of a high level of OXPHOS places cells under increased oxidative stress [10,11,12,13,15]. Evidence indicates that AMPK is activated in glucose-deprived cells by an increase of ROS [17,20] This appears to make sense in that, in the presence of high-level ROS and, thereby, the heavy accumulation of oxidatively damaged adducts, cells need to accelerate autophagy to facilitate damage disposition. Consistent with this idea, an impairment of the flux of basal autophagy and of those induced by other stimuli upon glucose deprivation has been reported [21]

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Results

Conclusion