Oxidative stress–dependent phosphorylation activates ZNRF1 to induce neuronal/axonal degeneration

Abstract

Oxidative stress induces EGFR-dependent phosphorylation and activation of ZNRF1 ubiquitin ligase in neurons, which promotes two major neurodegenerative pathways, apoptosis and axonal degeneration.

NADPH oxidases promote apoptosis by activating ZNRF1 ubiquitin ligase in neurons treated with an exogenously applied oxidant

“…We recently showed that ROS generated by NADPH oxidases serve as an intracellular signaling mediator after axonal injury in order to promote Wallerian degeneration by inducing EGFR-dependent ZNRF1 phosphorylation. 8 These findings combined with the present results suggest that the NADPH oxidase family of molecules play a more significant role in initiating neuronal degeneration than previously expected. NADPH oxidases accelerate apoptosis even when cell death is induced by an exogenously applied oxidant, suggesting that ROS generated by NADPH oxidases, which are closely located to the protein machineries that promote apoptosis, including those of ZNRF1 and EGFR, are more relevant for initiating neuronal degeneration than ROS generated elsewhere.…”

“…They were then washed, cultured in Neuro-medium containing 2% Neuro-Brew-21 and 1 mM glutamine for 24 h, and used in immunoblotting or immunostaining experiments. The antibodies used were as follows: anti-ZNRF1 antiserum; 7,10 antiserum for phosphorylated ZNRF1 at Y103; 8 rabbit anti-caspase 3 and anti-cleaved caspase 3 antibodies (9662, 9661, Cell Signaling Technology); rabbit polyclonal anti-βIII-tubulin antibody (Poly18020, BioLegend); anti-β-actin antibody (622101, BioLegend).

Figure 1.

…”

“…Interestingly, we found that downregulation of NOX3, 4, and DUOX2 are similarly effective in inhibiting 6OHDA-induced apoptosis, but not as effective as downregulation of ZNRF1-AKT-GSK3B signaling that we previously reported. 8 These results may suggest that more than 2 functionally redundant NADPH family molecules are located in the close proximity of the protein complex for promoting apoptosis. Elucidating the mechanisms underlying the activation of NADPH oxidases in neurons in more detail will provide an insight into the initiation mechanism of neurodegeneration.…”

“… 7-9 We found that, in response to increases in ROS, the epidermal growth factor receptor (EGFR)-dependent phosphorylation of ZNRF1 at the 103 rd tyrosine residue activated the ubiquitin ligase activity of ZNRF1 in order to target AKT for proteasomal degradation. 8 In a recent study, we employed a brain ischemia model (middle cerebral artery occlusion) and oxidant-induced Parkinson's disease model as models of ROS-induced neuronal cell death in vivo . 8 While the presence/production of excess ROS due to the significant changes in energy metabolism caused by ischemia is easily estimated in the brain ischemia model, previous studies have shown that the inhibition of NADPH oxidases significantly reduces neuronal damage, suggesting that NADPH oxidase-dependent ROS production beneath the neuronal membrane induced by ROS produced elsewhere plays a significant role in causing neuronal death in this model.…”

“… 8 In a recent study, we employed a brain ischemia model (middle cerebral artery occlusion) and oxidant-induced Parkinson's disease model as models of ROS-induced neuronal cell death in vivo . 8 While the presence/production of excess ROS due to the significant changes in energy metabolism caused by ischemia is easily estimated in the brain ischemia model, previous studies have shown that the inhibition of NADPH oxidases significantly reduces neuronal damage, suggesting that NADPH oxidase-dependent ROS production beneath the neuronal membrane induced by ROS produced elsewhere plays a significant role in causing neuronal death in this model. This finding prompted us to examine the contribution of NADPH oxidase activity in the oxidant-induced neuronal apoptosis model employed in our recent study.…”