Loss of BiP in hyperoxia does not activate the classic ER stress response. A549 cells were exposed to room air or hyperoxia for 24, 36, 48, and 72 hours for XBP1, PERK, and ATF6 assays. For XBP1 and PERK assays cells were treated with 0.5ug/ml tunicamycin (TM) or DMSO vehicle control for 24 hours. For the ATF6 assay cells were treated with 0.5ug/ml tunicamycin (TM) or DMSO vehicle control for 4 hours. (A) RNA was collected and RT-PCR for XBP1 was performed (u=unspliced XBP1, s=spliced XBP1). Whole cell lysates were immunoblotted for (B) phospho-PERK and (C) full length ATF6. (D) ATF6 protein levels were quantified, normalized to actin, and graphed. Data in panels A-C represent similar results from three independent experiments.