GCN2 activation by urea protects cells from osmotic stress

Abstract

General control non‐derepressible‐2 (GCN2) is one kinase known to be involved in the integrated stress response (ISR), and GCN2 activation (via phosphorylation) occurs in response to a variety of environmental stresses. GCN2 then phosphorylates eIF2α, a key step in ISR. Renal inner medullary cells are constantly exposed to high and variable concentrations of urea as a result of the urinary concentrating mechanism. To determine whether high concentrations of urea cause ISR in medullary cells, mIMCD3 cells were subjected to 250, 300 or 400 mM urea. GCN2 was rapidly activated by urea in a time‐dependent and dose‐dependent manner. At 250‐300mM urea GNC2 activation was reversible, however, when exposed to 400mM urea GCN2 activation persisted (~16 hours). eIF2α phosphorylation was also dose‐dependent. To investigate the effect of GCN2 activity on osmotic tolerance, GCN2 ‐/‐ and +/+ MEF cells were used. GCN2 ‐/‐ MEF cells were more susceptible to osmotic stress; 77% ± 5.0 of GCN2 +/+ MEF cells survived 300mM urea; in contrast, only 55% ± 9.6 of GCN2 ‐/‐ survived (P<0.05). We conclude that GCN2 is activated by urea and contributes to renal medullary cells' ability to survive osmotic stress.