Accumulation of unfolded proteins in the endoplasmic reticulum (ER) activates the unfolded protein response (UPR). In mammalian cells, UPR signals generated by several ER membrane resident proteins, including the bifunctional protein kinase endoribonuclease IRE1α, control cell survival and the decision to execute apoptosis. Processing of XBP1 mRNA by the RNase domain of IRE1α promotes survival of ER stress, while activation of the mitogen-activated protein kinase JNK by IRE1α late in the ER stress response promotes apoptosis. Here we show that activation of JNK in the ER stress response precedes activation of XBP1. This activation of JNK is dependent on IRE1α and TRAF2 and coincides with JNK-dependent induction of expression of several antiapoptotic genes, including cIAP1, cIAP2, XIAP, and BIRC6. ER-stressed jnk1-/- jnk2-/- mouse embryonic fibroblasts (MEFs) display more pronounced mitochondrial permeability transition and increased caspase 3/7 activity compared to wild type MEFs. Caspase 3/7 activity is also elevated in ER-stressed ciap1-/- ciap2-/-, and xiap-/- MEFs. These observations suggest that JNK-dependent transcriptional induction of several inhibitors of apoptosis contributes to inhibiting apoptosis early in the ER stress response.

Accumulation of unfolded proteins in the endoplasmic reticulum (ER) causes ER stress and activates a signalling network known as the unfolded protein response (UPR). Here we characterise how ER stress and the UPR inhibit insulin signalling. We find that ER stress inhibits insulin signalling by depleting the cell surface population of the insulin receptor. ER stress inhibits proteolytic maturation of insulin proreceptors by interfering with transport of newly synthesised insulin proreceptors from the ER to the plasma membrane. Activation of AKT, a major target of the insulin signalling pathway, by a cytosolic, membrane-bound chimera between the AP20187-inducible FV2E dimerisation domain and the cytosolic protein tyrosine kinase domain of the insulin receptor was not affected by ER stress. Hence, signalling events in the UPR, such as activation of the JNK MAP kinases or the pseudokinase TRB3 by the ER stress sensors IRE1α and PERK, do not contribute to inhibition of signal transduction in the insulin signalling pathway. Indeed, pharmacologic inhibition and genetic ablation of JNKs, as well as silencing of expression of TRB3, did not restore insulin sensitivity or rescue processing of newly synthesised insulin receptors in ER-stressed cells.

Obesity is associated with endoplasmic reticulum (ER) stress and activation of the unfolded protein response (UPR) in adipose tissue. In this study we identify physiological triggers of ER stress and of the UPR in adipocytes in vitro. We show that two markers of adipose tissue remodelling in obesity, glucose starvation and hypoxia, cause ER stress in 3T3-F442A and 3T3-L1 adipocytes. Both conditions induced molecular markers of the IRE1a and PERK branches of the UPR, such as splicing of XBP1 mRNA and CHOP, as well as transcription of the ER stress responsive gene BiP. Hypoxia also induced an increase in phosphorylation of the PERK substrate eIF2a. By contrast, physiological triggers of ER stress in many other cell types, such as the saturated fatty acid palmitic acid, cholesterol, or several inflammatory cytokines including TNF-a, IL-1b, and IL-6, do not cause ER stress in 3T3-F442A and 3T3-L1 adipocytes. Our data suggest that physiological changes associated with remodelling of adipose tissue in obesity, such as hypoxia and glucose starvation, are more likely physiological ER stressors of adipocytes than the lipid overload or hyperinsulinemia associated with obesity.