Abstract
The integrated stress response is characterized by the phosphorylation of eukaryotic initiation factor-2α (eIF2α) on serine 51 by one out of four specific kinases (EIF2AK1 to 4). Here we provide three series of evidence suggesting that macroautophagy (to which we refer to as autophagy) induced by a variety of distinct pharmacological agents generally requires this phosphorylation event. First, the induction of autophagic puncta by various distinct compounds was accompanied by eIF2α phosphorylation on serine 51. Second, the modulation of autophagy by >30 chemically unrelated agents was partially inhibited in cells expressing a non-phosphorylable (S51A) mutant of eIF2α or lacking all four eIF2α kinases, although distinct kinases were involved in the response to different autophagy inducers. Third, inhibition of eIF2α phosphatases was sufficient to stimulate autophagy. In synthesis, it appears that eIF2α phosphorylation is a central event for the stimulation of autophagy.
Highlights
The so-called integrated stress response[1,2,3] is characterized by the phosphorylation of eukaryotic initiation factor2α
Correlation of peIF2α and autophagy As a first approach to investigate the interdependence between autophagy and peIF2α, we measured the phosphorylation of eukaryotic initiation factor2α (eIF2α) in human osteosarcoma U2OS cells by means of an immunofluorescence staining protocol with a phosphoneoepitope-specific antibody recognizing eIF2α phosphorylated on serine 51
There are three major arguments that favor the hypothesis that peIF2α is important for the activation of autophagy: (i) the stimulation of autophagic puncta and peIF2α correlates for many autophagy modulators; (ii) cells bearing a non-phosphorylable eIF2α mutant or lacking all known eIF2α kinases are refractory to autophagy induction by most stimuli; and (iii) activation of peIF2α by inhibition of a specific set of phosphatases suffices to trigger autophagy
Summary
The so-called integrated stress response[1,2,3] is characterized by the phosphorylation of eukaryotic initiation factor2α (eIF2α). The phosphorylation of eIF2α occurs on serine 51 in response to the activation of one out of four eIF2α kinases[4,5,6]. EIF2AK1, commonly known as heme-regulated inhibitor (HRI), is activated by oxidative, osmotic, and heat stress, as well as by arsenic and redaporfin-mediated photodynamic therapy[7,8,9,10]. EIF2AK2, commonly known as protein kinase R (PKR), is activated by viruses and alcohol[11,12]. EIF2AK3, commonly known as protein kinase Rlike endoplasmic reticulum (ER) kinase (PERK), is activated by ER stress, because unfolded proteins in the ER lumen occupy the chaperone GRP78, which releases PERK from inhibition[13,14]. A variety of rather distinct stressors converge on eIF2α phosphorylation (peIF2α)
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