Figure 8. Signaling of p53 expression by NF-kB during endoplasmic reticulum stress. Accumulation of unfolded or misfolded proteins in the endoplasmic reticulum induces activation of multiple signaling pathways. Many studies have indicated that activation of IRE1, ATF6, and PERK represents the standard UPR pathways and turns on the expression of many downstream genes such as GRP78. ER stress induced caspase-12/caspase-9/caspase-3, PERK/ATF- 4/CHOP, IRE1/Ask1/JNK, and PERK/eIF2a/NF-kB signaling pathways and then those pathways may alter cellular homeostasis by regulating multiple genes, resulting in regulation of cell death or survival. This pathway (indicated by heavy arrows) is demonstrated in this report. We demonstrated that induction of p53 expression is mediated by NF-kB during ER stress. doi:10.1371/journal.pone.0039120.g008 I and Topo II inhibitors in relation to ATM and Chk2 activation [44]. In vitro evidence suggests that DNA damage-induced p53 Ser15 phosphorylation can block the interaction of p53 with its inhibitor, MDM2, and stimulate association with transcriptional co-activator proteins such as p300 [31]. Phosphorylation of p53 on Ser 15 and Ser37 sites can be regulated by ATM, ATR and DNA- PK, thus inhibiting the ubiquitin degradation of p53, promoting p53 activation and accumulation [45]. Therefore, our finding of p53 phosphorylation on Ser15 by ER stress shows these processes may participate in the regulation of ER stress-induced cell death. However, it is unclear what role Ser15 phosphorylation plays in ER stress processes and how this modification affects context- dependent programs of p53-regulated gene expression and their biological consequences. The relationship between p53 phosphor- ylation on Ser15 site and ER stress-induced cell death warrants further investigation. Previous study indicated that regulation of NF-kB signaling pathway plays an important role during ER stress. From our previous results, we demonstrated that induction of COX-2 expression was mediated through NF-kB and p38 pathways in response to ER stress [17]. The role of NF-kB in COX-2 expression may suggest that NF-kB plays anti-apoptosis effects in response to ER stress. In addition to NF-kB and COX-2, ER stress can enhance proinflammatory NF-kB activation via C/EBP Homologous Protein (CHOP) and maintain an increased level of IL-8 production in human intestinal epithelial cells [46]. There is modulation of cell death in heart failure by NF-kB, which NF-kB activation required for ER stress-mediated apoptosis, whereas References 1. Kaufman RJ (2002) Orchestrating the unfolded protein response in health and disease. J Clin Invest 110(10): 1389–1398. 2. Paschen W, Frandsen A (2001) Endoplasmic reticulum dysfunction – a common denominator for cell injury in acute and degenerative diseases of the brain? J Neurochem 79(4): 719–725. 3. Shi Y, Taylor SI, Tan SL, Sonenberg N (2003) When translation meets metabolism: multiple links to diabetes. Endocr Rev 24(1): 91–101. Regulation of p53 Expression by NF-kB in ER Stress abrogation of myocyte NF-kB shifted the ER stress response to one of adaptation and survival [47]. These results implicated, in response to ER stress, ER-initiated pathways signal alarm by activating NF-kB transcription factor that induces expression of genes encoding mediation of host defense. Activation of both the PERK and IRE1 pathways leads to regulation of the NF-kB-IKK signaling pathway during ER stress through activation of IKK or degradation of the p65 subunit. The ATF6 branch can also regulate NF-kB activity [48]. All of these signals contribute to the triggering of apoptotic responses when ER stress is excessive, prolonged, or insufficiently neutralized, and all UPR sensors are activated and lead to the induction of both pro-apoptotic and anti- apoptotic factors. Previous studies have shown that multiple pathways are involved in ER stress-induced apoptosis, such as caspase12, PERk-eIF2a, IRE-1-JNK, and p53 pathways. Persis- tent ER stress can also trigger a switch in the UPR signal pathways from prosurvival to proapoptoic pathways, such as PERk-eIF2a and IRE-1-JNK pathways. On the other hand, many studies have also indicated that three phases of adaptation, alarm, and apoptosis mechanisms are involved in regulating prolonged ER stress [49]. When adaptation fails, incessant ER stress can trigger the signal alarm by activation of NF-kB. In PERk-eIF2a pathway, PERK protein activates its intrinsic kinase activity, resulting in phosphorylation of eIF2a and induction of NF-kB activity. For IRE-1-TRAF2 pathway, Ire1 binds TRAF2, signaling down- stream kinases activation of NF-kB and c-jun, causing expression of genes associated with alarm and proapoptotic phases. Based on those research findings, NF-kB appears to play an important role in regulating signal pathways associated with ER stress-induced cell survival and apoptosis. Because during ER stress, ER-initiated pathways signal alarm by activation of NF-kB a transcription factor that induces expression of genes encoding mediators of host defense such as COX-2. However, Excessive and prolonged ER stress, activation of NF-kB turns to regulate cell death-associated genes expression such as p53, resulting in increase cell apoptosis. In addition, previous study indicated that increased level of ER stress which sensitizes drugs-resistant cells, and combination of ER stress-inducing agents is a novel therapeutic strategy for cancer cells [50,51]. Our data provides a molecular basis for consider- ation of employing ER stress in killing of cancer cells directly or sensitizing the cells to other cytotoxic chemotherapy. In summary, ER stress causes cell death by increasing the expression of p53 and activating NF-kB which is involved in the enhanced expression of p53. Previous studies have indicated that chemotherapeutic drugs cause cell death through ER stress, such as with Cisplatin [52]. These results may provide an important therapeutic strategy for chemotherapeutic drugs through ER stress-associated signaling pathways. Author Contributions Conceived and designed the experiments: W-CL Y-CC Y-SC M-DL CCCW K-HL J-HH. Performed the experiments: W-CL Y-CC Y-SC J- HH. Analyzed the data: W-CL Y-CC J-HH. Contributed reagents/ materials/analysis tools: M-DL CCCW K-HL Y-NT I-JS J-HH. Wrote the paper: J-HH. 4. Ozcan U, Cao Q, Yilmaz E, Lee AH, Iwakoshi NN, et al. (2004) Endoplasmic reticulum stress links obesity, insulin action, and type 2 diabetes. Science 306(5695): 457–461. 5. Calfon M, Zeng H, Urano F, Till JH, Hubbard SR, et al. (2002) IRE1 couples endoplasmic reticulum load to secretory capacity by processing the XBP-1 mRNA. Nature 415(6867): 92–96. PLoS ONE | www.plosone.org 10 July 2012 | Volume 7 | Issue 7 | e39120