Hyperoxia-induced activation of the integrated stress response in the newborn rat lung

Diverse environmental stresses stimulate eukaryotic translation initiation factor 2α (eIF2α) phosphorylation, leading to a stress-resistant state characterized by global attenuation of protein synthesis and induction of cytoprotective genes. The signal transduction network culminating in these effects is referred to as the integrated stress response (ISR) or, when initiated by misfolded proteins within the endoplasmic reticulum (ER), the unfolded protein response (UPR). Given that we previously reported that exposure of 4-day-old Sprague-Dawley rats to 95% O(2) (Ox) diminishes global pulmonary protein synthesis and increases eIF2α phosphorylation, we conducted the current study to determine whether Ox activates the ISR or UPR. We found that Ox-induced alterations in ER morphology of alveolar type II cells and interstitial fibroblasts were not associated with activation of the UPR sensors PERK or activating transcription factor (ATF) 6 or with X-box binding protein-1 mRNA splicing in whole lung extracts. Exposure to Ox enhanced ATF4 immunoreactivity and nuclear protein content, followed by a 2- and 5-fold increase in ATF3 protein and mRNA expression, respectively. The accumulation of nuclear ATF4 protein coincided with induction of glutamate-cysteine ligase catalytic subunit, an ISR-responsive gene. Immunohistochemistry revealed that changes in ATF3/4 expression were prominent in the alveolus, whereas primary cell culture implicated epithelial and endothelial cells as targets. Finally, induction of ISR intermediates in the intact lung occurred in the absence of the phosphorylation of PKR, JNK, ERK1/2, and p38 MAPK. These findings demonstrate that Ox activates the ISR within the newborn lung and highlight regional and cell-specific alterations in the expression ISR transcription factors that regulate redox balance.