Ribose 5-phosphate isomerase inhibits LC3 processing and basal autophagy

Jacob Heintze,Joana R Costa,Melanie Weber,Robin Ketteler

doi:10.1016/j.cellsig.2016.06.015

Jacob Heintze, Joana R Costa + Show 2 more

Open Access

https://doi.org/10.1016/j.cellsig.2016.06.015

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Abstract

Autophagy and cellular metabolism are tightly linked processes, but how individual metabolic enzymes regulate the process of autophagy is not well understood. This study implicates ribose-5-phosphate isomerase (RPIA), a key regulator of the pentose phosphate pathway, in the control of autophagy. We used a dual gene deletion strategy, combining shRNA-mediated knockdown studies with CRISPR/Cas9 genome editing. Knockdown of RPIA by shRNA or genomic deletion by CRISPR/Cas9 genome editing, results in an increase of ATG4B-mediated LC3 processing and in the appearance of LC3-positive autophagosomes in cells. Increased LC3 processing upon knockdown of RPIA can be reversed by treatment with the antioxidant N-acetyl cysteine. The results are consistent with a model in which RPIA suppresses autophagy and LC3 processing by modulation of redox signaling.

Highlights

Macroautophagy is a physiological process in response to low levels of metabolites and nutrients, that provides additional energy to the cell through the induction of a lysosomal degradation pathway and recycling of cellular constituents [31]
Upon transfection of shRNAs targeting ribose-5-phosphate isomerase (RPIA), we observed a significant increase in LC3-II compared to LC3-I, suggesting an increase in basal autophagy (Fig. 1A), with all four shRNA sequences being effective to cause an increase in LC3-II to LC3-I and LC3-II to loading control ratios (Fig. 1B, C)
We investigated whether the effect of RPIA depletion on LC3 processing is via the autophagy protease ATG4B

Summary

Introduction

Macroautophagy (hereafter autophagy) is a physiological process in response to low levels of metabolites and nutrients, that provides additional energy to the cell through the induction of a lysosomal degradation pathway and recycling of cellular constituents [31]. Conditions such as starvation, stress and pathogen infections induce autophagy, and basal autophagy is an important process for cellular homeostasis. In the process of autophagosome maturation, members of the LC3 family are cleaved by ATG4 proteases, lipidated and incorporated into maturing autophagosomes, in which LC3B (hereafter LC3) is the predominant form. The various stages of LC3 maturation can be measured by immunoblotting

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Conclusion