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Logo of nihpaAbout Author manuscriptsSubmit a manuscriptHHS Public Access; Author Manuscript; Accepted for publication in peer reviewed journal;
 
Cell Cycle. Author manuscript; available in PMC 2011 January 5.
Published in final edited form as:
PMCID: PMC2834424
NIHMSID: NIHMS177789

The roles of nitric oxide synthase and eIF2alpha kinases in regulation of cell cycle upon UVB-irradiation

Lei Wang,1,2 Yan Liu,1,2 and Shiyong Wu1,2,3,*

Abstract

In response to ultraviolet light (UV)-induced damage, cells initiate cellular recovery mechanisms including activation of repair genes and redistribution of cell cycle phases. While most studies have focused on DNA damage-inducible transcriptional regulation of cell cycle checkpoints, translational regulation also plays an important role in control of cell cycle progression upon UV-irradiation. UV-irradiation activates two kinases, PERK and GCN2, which phosphorylate the alpha subunit of eukaryotic initiation factor 2 (eIF2α) and subsequently inhibit protein synthesis. We recently identified an upstream regulator, nitric oxide synthase (NOS), which controls the activation of both PERK and GCN2 upon UVB-irradiation. Our data suggested that UVB induces NOS activation and NO production, which reacts with superoxide (O2 •−) to form peroxynitrite (ONOO) and activate PERK. The NO production also leads to L-Arg depletion and GCN2 activation. The elevation of nitric oxide and activation of PERK/GCN2 have been shown to play roles in regulation of cell cycle upon UVB irradiation. In the present study, we show that the cell cycle phases were redistributed by inhibition of NOS activation or reduction of oxidative stress upon UVB irradiation, indicating the roles of NO and its oxidative products in regulation of cell cycle. We also demonstrate that both PERK and GCN2 were involved in regulation of cell cycle upon UVB-irradiation, but the regulation is independent of eIF2α phosphorylation. While the mechanism for UVB-induced cell cycle control is yet to be unraveled, we here discuss the differential roles of NOS, PERK and GCN2 in regulation of cell cycle upon UVB-irradiation.

Keywords: nitric oxide, peroxynitrite, cell cycle, translation regulation, EIF2AK, eIF2α phosphorylation

Cells respond to ultraviolet light (UV) by reprogramming signaling circuits that control cellular physiology, including cell cycle regulation and apoptotic cell death.15 The roles of transcriptional regulation in UV-induced cell cycle arrest and apoptosis are intensively studied.611 In the latest decade, the impacts of translation, especially the phosphorylation on the α subunit of translation initiation factor 2 (eIF2α), on UV-induced signaling circuits, are being elucidated.1220 The phosphorylation eIF2α is a universal response of eukaryotic cells to various types of stress, and is regulated by different eIF2α kinases.2123 Two eIF2α kinases (EIF2AK), the dsRNA-dependent protein kinase-like ER kinase (PERK, EIF2AK3) and the general control nonderepressible protein kinase 2 (GCN2, EIF2AK4), were shown to be responsible for UV-induced phosphorylation of eIF2α.16,18 PERK is an ER membrane localized kinase, which is activated by endoplasmic reticulum (ER)-stress.24,25 GCN2 is an amino acid abundance controlled eIF2α kinase, which is activated during amino acid starvation.26,27 While the roles of PERK and GCN2 in UV-induced translation inhibition and downstream signal transduction are elucidated,14,15,20,28 their key upstream activator(s) have not been identified.

In a recent paper published in the September issue of The Journal of Biological Chemistry, we provided evidence that UVB activates constitutive nitric oxide synthase (cNOS), which coordinately regulates the activation of both PERK/GCN2 and sequentially the phosphorylation of eIF2α in human keratinocytes HaCaT.29 We demonstrated that UVB-activated NOS rapidly generates NO from L-Arg, which had a concentration of less than 2.5 µM in HaCaT cells. Our data suggested that the consumption of L-Arg led to a shortage of the amino acid, which activates GCN2. The UVB-induced elevation of superoxide (O2•−)30 and depletion of L-Arg led to cNOS uncoupling and generation of peroxynitrite (ONOO), which is a strong oxidant3133 that can be rapidly produced from NO and O2•−.32,33 The accumulation of intracellular ONOO induces ER-stress and PERK activation.34 In addition to keratinocytes, we also studied the regulation of eIF2α phosphorylation in wild-type mouse embryonic fibroblasts (MEF), PERK knockout (MEFPERK−/−), GCN2 knockout (MEFGCN2−/−) cells and the eIF2α Ser51 Ala mutant (MEFA/A) cells. Distinctive from HaCaT cells, all three MEF cell lines have higher background phosphorylation of eIF2α, which was maintained by basal NOS activity and oxidative stress. L-Arg-shortage-mediated GCN2 activation appears to play a more significant role for maintaining basal eIF2α phosphorylation in MEFPERK−/− cells. However, UVB-induced eIF2α phosphorylation in MEFPERK−/− cells mainly resulted from oxidative-stress. In MEFGCN2−/− cells, L-Arg biosynthesis is downregulated due to the lower activity of GCN4, whose activity is positively controlled by GCN2.35,36 The down-regulation of L-Arg biosynthesis leads to L-Arg depletion and sequentially to NOS uncoupling and higher oxidative-stress levels, which prevented UVB from further inducing eIF2α phosphorylation. Our results demonstrated that a chronic elimination of a gene could rearrange the entire signaling circuit in response to a stimuli and that interpretation from data generated from knockout cells could be misleading. In the following chapters, we analyzed cell cycles of HaCaT and MEF cells in response to UVB-irradiation. We demonstrated that NOS, PERK and GCN2 play differential roles in regulation of cell cycle upon UVB-irradiation.

NOS Activation in Combination with Oxidative Stress Regulates G1/S Phase Cell Cycle Arrest in Keratinocytes after UVB-Irradiation

NO regulates a wide range of cell functions such as cell cycle,37,38 cell proliferation3941 and apoptosis.41 However, the role of NO in cell cycle control is controversial. On one hand, it shows that exposure to NO derived from either endogenous NOS or exogenous NO donors blocks cell cycle progression on G0/G1 or G2/M phase depending on cell types.4244 TNFα, IFNγ and IL-1β are commonly used as iNOS stimulators to produce endogenous NO, which blocks the cell’s entry to the G0/G1 phase.45 NO donors also induce G1 arrest in different cell types such as smooth muscle cells,46 breast cancer cells,47 human pro-monocytic cells U937,48 and melanoma cells.49 On the other hand, some research also demonstrated that exogenous NO generated from S-nitrosoglutathione prevents gamma-irradiation-induced G1 arrest by impairing p53 conformation.50

To determine the role of activation NOS in the regulation of cell cycle upon UVB-irradiation, we pretreated the cells with a broad NOS inhibitor, a N-substituted L-arginine analog LNMMA (100 µM) for 1 h before and 4 h after UVB-irradiation (50 mJ/cm2). At 24 h post-irradiation, the cell cycle was analyzed by flow cytometry. Our data showed that the G1 phase fraction increases from 49 ± 3% to 70 ± 2% after UVB irradiation (Fig. 1). With treatment of LNMMA, the amount of cells in G1 phase was increased to 64 ± 4% after UVB-irradiation (Fig. 1). With the consideration of the 4% increase of G1 phase cells by treating with LNMMA alone, the results suggested that NOS plays a role in regulation of G1 phase arrest upon UVB. Besides NO, UV also induces an elevation of ONOO, which is a stronger oxidant with a biological half-life near 100 ms.51 To determined the role of ONOO in regulation of cell cycle, we use a glutathione (GSH) synthesis precursor N-acetyl-L-cysteine (LNAC), which was reported to specifically reduce intracellular ONOO levels in multiple cell types.52,53 Our data showed that treating the cells with LNAC significantly attenuates UVB-induced G1 arrest cell proportion from 70 ± 2% to 50 ± 1% (Fig. 1), which is similar to the control without UVB-irradiation (Fig. 1). This result implies that ONOO plays a key role in UVB-induced keratinocytes G1 arrest, and the role of NO in cell cycle control might mediated by formation of ONOO in stead of itself.

Figure 1
NOS and ONOO mediate UVB-induced cell cycle G1 arrest and S phase change in keratinocytes. HaCat cells were treated with 100 µM LNMMA or 25 mM LNAC for 1 hour and then UVB-irradiated (50 mJ/cm2). Immediately after irradiation, the cells ...

In addition, the NO/ONOO-mediated G1 arrest is accompanied with a reduced S phase, which represents the DNA synthesis phase in the cell cycle, and usually used as a proliferative index for cell growth. Our data showed that after UVB-irradiation, the percentage of cells in S phase dropped from 33 ± 2% to 6.8 ± 2% (Fig. 1), exhibiting a significant halt in DNA synthesis caused by UVB insults. Inhibition of NOS activity by LNMMA recovered DNA synthesis from UVB-irradiation, the cells in S phase increased from 7 ± 2% to 16 ± 4%. Reduction of ONOO by LNAC increased the cells in S phase from 7 ± 2% to 30 ± 2% in UVB treatment. These results suggested that NOS activation in combination with oxidative stress regulates G1/S phase cell cycle in keratinocytes upon UVB-irradiation.

The Roles of GCN2, PERK and eIF2α in Regulation of Cell Cycle upon UVB Irradiation

Both PERK and GCN2 regulate cell cycle and apoptosis in response to various stimuli.14,19,28,5458 ER-stress-induced PERK activation or overexpression of PERK downregulates cell cycle regulator cyclin D1 and the tumor suppressor p53, leading to G1 arrest.56,59 Overexpression a dominant negative truncated PERK lacking its kinase domain prevented unfolding protein response (UPR)-induced cyclin D1 loss and cell cycle arrest.54 GCN2 activation accompanied with increased eIF2α phosphorylation controls the progression of the G1 phase and delays entry to S phase with fission yeast cells in response to UV-irradiation.28,60

In our article published in the September 4th JBC, we demonstrated that NOS and oxidative stress mediate the activation of PERK/GCN2 and phosphorylation of eIF2α.29 To determine whether the UVB-induced activation of the EIF2AKs and phosphorylation of eIF2α also play a role in the regulation of cell cycle, we analyzed the cell cycle patterns of a series of MEF cell lines after UVB irradiation. These cell lines included: wild-type (MEFWT), PERK knockout (MEFPERK−/−), GCN2 knockout (MEFGCN2−/−) and the eIF2α Ser51 Ala mutant (MEFA/A). Our data showed that MEFWT cells behaved very much different from HaCaT cells. Without UVB-treatment, MEFWT cells have relatively fewer cells in G1/G2 phases and more cells in S phase than HaCaT cells (Fig. 2). Knocking out PERK or GCN2 increased 16.1% or 7.7% of the cells in G1 phase respectively, decreased 16.9% or 8.5% of the cells in S phase respectively, but had almost no impact on G2 phase (Fig. 2). In contrast, total elimination of eIF2α phosphorylation by knocking in a non-phosphorylatable eIF2α (S51A) mutant (MEFA/A) reduced 5.6% and 7.7% of the cells in the G1 and S phases, while an increase of 3.8% was observed in the G2 phase. However, the changes in MEFA/A cell cycle are not statistically significant to wild type MEF cells (Fig. 2). These results suggested that basal PERK/GCN2 activities are required for maintaining an efficient G1 checkpoint. However, it is not clear whether PERK or GCN2 regulates cell cycle via eIF2α phosphorylation since elimination of the phosphorylation had no significant effects on the patterns of cell cycle. After UVB-irradiation, there was a shift of 4.4% of MEFWT cells from the G1 phase to the G2 phase with no change observed at the S phase (Fig. 2). This result was very different from that of the HaCaT cells after UVB-irradiation, which showed a significantly increased portion of cells in the G1/G2 phases and a reduced portion of the cells in the S phase. For MEFPERK−/− or MEFGCN2−/− cells, UVB-irradiation shifted approximately 10% of cells from the G1 phase to the G2 phase (Fig. 2). This data indicated that PERK and GCN2 both played a role in regulation of the cell cycle in response to UV-induced DNA damage and oxidative stress in mammalian cells, as previously reported that GCN2 did in yeast.28,61 Interestingly, while reduction of PERK or GCN2 activity promoted UVB-induced cell cycle shift from the G1 phase to the G2 phase, elimination of eIF2α phosphorylation prevented UVB-induced cell cycle shift (Fig. 2). These results suggested that PERK and GCN2 might regulate the cell cycle in mammalian cells via an eIF2α phosphorylation independent pathway as GCN2 does in yeast.19 The eIF2α phosphorylation-independent regulation of cell cycle by EIF2AKs could be mediated via promotion of p53 degradation,62 which plays a key role in regulation of cell cycle upon UVB-irradiation.63

Figure 2
PERK and GCN2 regulates cell cycle G2 arrest upon UVB irradiation in MEF cells. MEF cells were irradiated by UVB (50 mJ/cm2). Twelve hours after UVB irradiation, the cells were fixed in 70% ethanol. The cell cycle was analyzed by propidium iodide (PI) ...

Acknowledgements

We thank Mr. Oliver Luke Carpenter and Ms. Molly Monica for reading the manuscript. This work was supported by National Institutes of Health Grant RO1 CA86928 (to S.W.) and R56 CA086928 (to S.W.)

References

1. Kurki S, Peltonen K, Laiho M. Nucleophosmin, HDM2 and p53: players in UV damage incited nucleolar stress response. Cell Cycle. 2004;3:976–979. [PubMed]
2. Fotedar R, Bendjennat M, Fotedar A. Role of p21WAF1 in the cellular response to UV. Cell Cycle. 2004;3:134–137. [PubMed]
3. Batista LF, Kaina B, Meneghini R, Menck CF. How DNA lesions are turned into powerful killing structures: insights from UV-induced apoptosis. Mutat Res. 2009;681:197–208. [PubMed]
4. Rundhaug JE, Fischer SM. Cyclo-oxygenase-2 plays a critical role in UV-induced skin carcinogenesis. Photochem Photobiol. 2008;84:322–329. [PubMed]
5. Soehnge H, Ouhtit A, Ananthaswamy ON. Mechanisms of induction of skin cancer by UV radiation. Front Biosci. 1997;2:538–551. [PubMed]
6. Westfall MD, Joyner AS, Barbieri CE, Livingstone M, Pietenpol JA. Ultraviolet radiation induces phosphorylation and ubiquitin-mediated degradation of DeltaNp63alpha. Cell Cycle. 2005;4:710–716. [PubMed]
7. Cooper SJ, Bowden GT. Ultraviolet B regulation of transcription factor families: roles of nuclear factor-kappa B (NFkappaB) and activator protein-1 (AP-1) in UVB-induced skin carcinogenesis. Curr Cancer Drug Targets. 2007;7:325–334. [PMC free article] [PubMed]
8. Ulm R, Nagy F. Signalling and gene regulation in response to ultraviolet light. Curr Opin Plant Biol. 2005;8:477–482. [PubMed]
9. Terui T, Okuyama R, Tagami H. Molecular events occurring behind ultraviolet-induced skin inflammation. Curr Opin Allergy Clin Immunol. 2001;1:461–467. [PubMed]
10. Fisher GJ, Voorhees JJ. Molecular mechanisms of photoaging and its prevention by retinoic acid: ultraviolet irradiation induces MAP kinase signal transduction cascades that induce Ap-1-regulated matrix metalloproteinases that degrade human skin in vivo. J Investig Dermatol Symp Proc. 1998;3:61–68. [PubMed]
11. Trautinger F, Kindas-Mugge I, Knobler RM, Honigsmann H. Stress proteins in the cellular response to ultraviolet radiation. J Photochem Photobiol B. 1996;35:141–148. [PubMed]
12. Laszlo CF, Fayad S, Carpenter OL, George KS, Lu W, Saad AA, et al. The role of translational regulation in ultraviolet C light-induced cyclooxygenase-2 expression. Life Sci. 2009;85:70–76. [PMC free article] [PubMed]
13. Laszlo CF, Wu S. Mechanism of UV-induced IkappaBalpha-independent activation of NFkappaB. Photochem Photobiol. 2008;84:1564–1568. [PMC free article] [PubMed]
14. Parker SH, Parker TA, George KS, Wu S. The roles of translation initiation regulation in ultraviolet light-induced apoptosis. Mol Cell Biochem. 2006;293:173–181. [PubMed]
15. Wu S, Tan M, Hu Y, Wang JL, Scheuner D, Kaufman RJ. Ultraviolet light activates NFkappaB through translational inhibition of IkappaBalpha synthesis. J Biol Chem. 2004;279:34898–34902. [PubMed]
16. Wu S, Hu Y, Wang JL, Chatterjee M, Shi Y, Kaufman RJ. Ultraviolet Light Inhibits Translation through Activation of the Unfolded Protein Response Kinase PERK in the Lumen of the Endoplasmic Reticulum. J Biol Chem. 2002;277:18077–18083. [PubMed]
17. Liu G, Zhang Y, Bode AM, Ma WY, Dong Z. Phosphorylation of 4E-BP1 is mediated by the p38/MSK1 pathway in response to UVB irradiation. J Biol Chem. 2002;277:8810–8816. [PubMed]
18. Deng J, Harding HP, Raught B, Gingras AC, Berlanga JJ, Scheuner D, et al. Activation of GCN2 in UV-irradiated cells inhibits translation. Curr Biol. 2002;12:1279–1286. [PubMed]
19. Marbach I, Licht R, Frohnmeyer H, Engelberg D. Gcn2 mediates Gcn4 activation in response to glucose stimulation or UV radiation not via GCN4 translation. J Biol Chem. 2001;276:16944–16951. [PubMed]
20. Jiang HY, Wek RC. GCN2 phosphorylation of eIF2alpha activates NFkappaB in response to UV irradiation. Biochem J. 2005;385:371–380. [PubMed]
21. Clemens MJ. Initiation factor eIF2alpha phosphorylation in stress responses and apoptosis. Prog Mol Subcell Biol. 2001;27:57–89. [PubMed]
22. Proud CG. eIF2 and the control of cell physiology. Semin Cell Dev Biol. 2005;16:3–12. [PubMed]
23. Kaufman RJ. Control of gene expression at the level of translation initiation. Curr Opin Biotechnol. 1994;5:550–557. [PubMed]
24. Harding HP, Zhang Y, Ron D. Protein translation and folding are coupled by an endoplasmic-reticulum-resident kinase. Nature. 1999;397:271–274. [PubMed]
25. Kaufman RJ. Stress signaling from the lumen of the endoplasmic reticulum: coordination of gene transcriptional and translational controls. Genes Dev. 1999;13:1211–1233. [PubMed]
26. Berlanga JJ, Santoyo J, De Haro C. Characterization of a mammalian homolog of the GCN2 eukaryotic initiation factor 2alpha kinase. Eur J Biochem. 1999;265:754–762. [PubMed]
27. Sood R, Porter AC, Olsen D, Cavener DR, Wek RC. A Mammalian Homologue of GCN2 Protein Kinase Important for Translational Control by Phosphorylation of Eukaryotic Initiation Factor-2alpha. Genetics. 2000;154:787–801. [PubMed]
28. Grallert B, Boye E. The Gcn2 kinase as a cell cycle regulator. Cell Cycle. 2007;6:2768–2772. [PubMed]
29. Lu W, Laszlo CF, Miao Z, Chen H, Wu S. The role of nitric oxide synthase in regulation of ultraviolet light-induced phosphorylation of the alpha-subunit of eukaryotic initiation factor 2. J Biol Chem. 2009 [PMC free article] [PubMed]
30. Wiswedel I, Keilhoff G, Dorner L, Navarro A, Bockelmann R, Bonnekoh B, et al. UVB irradiation-induced impairment of keratinocytes and adaptive responses to oxidative stress. Free Radic Res. 2007;41:1017–1027. [PubMed]
31. Pou S, Keaton L, Surichamorn W, Rosen GM. Mechanism of superoxide generation by neuronal nitric-oxide synthase. J Biol Chem. 1999;274:9573–9580. [PubMed]
32. Groves JT. Peroxynitrite: reactive, invasive and enigmatic. Curr Opin Chem Biol. 1999;3:226–235. [PubMed]
33. Beckman JS, Koppenol WH. Nitric oxide, superoxide and peroxynitrite: the good, the bad and ugly. Am J Physiol. 1996;271:1424–1437. [PubMed]
34. Dickhout JG, Hossain GS, Pozza LM, Zhou J, Lhotak S, Austin RC. Peroxynitrite causes endoplasmic reticulum stress and apoptosis in human vascular endothelium: implications in atherogenesis. Arterioscler Thromb Vasc Biol. 2005;25:2623–2629. [PubMed]
35. Hinnebusch AG. Novel mechanisms of translational control in Saccharomyces cerevisiae. Trends Genet. 1988;4:169–174. [PubMed]
36. Crabeel M, Huygen R, Verschueren K, Messenguy F, Tinel K, Cunin R, et al. General amino acid control and specific arginine repression in Saccharomyces cerevisiae: physical study of the bifunctional regulatory region of the ARG3 gene. Mol Cell Biol. 1985;5:3139–3148. [PMC free article] [PubMed]
37. Zhang J, Wang S, Kern S, Cui X, Danner RL. Nitric oxide downregulates polo-like kinase 1 through a proximal promoter cell cycle gene homology region. J Biol Chem. 2007;282:1003–1009. [PubMed]
38. Lu Q, Jourd’Heuil FL, Jourd’Heuil D. Redox control of G(1)/S cell cycle regulators during nitric oxide-mediated cell cycle arrest. J Cell Physiol. 2007;212:827–839. [PubMed]
39. Janssens S, Flaherty D, Nong Z, Varenne O, van Pelt N, Haustermans C, et al. Human endothelial nitric oxide synthase gene transfer inhibits vascular smooth muscle cell proliferation and neointima formation after balloon injury in rats. Circulation. 1998;97:1274–1281. [PubMed]
40. Ignarro LJ, Buga GM, Wei LH, Bauer PM, Wu G, del Soldato P. Role of the arginine-nitric oxide pathway in the regulation of vascular smooth muscle cell proliferation. Proc Natl Acad Sci USA. 2001;98:4202–4208. [PubMed]
41. Brune B. Nitric oxide: NO apoptosis or turning it ON? Cell Death Differ. 2003;10:864–869. [PubMed]
42. Sarkar R, Gordon D, Stanley JC, Webb RC. Cell cycle effects of nitric oxide on vascular smooth muscle cells. Am J Physiol. 1997;272:1810–1818. [PubMed]
43. Tanner FC, Meier P, Greutert H, Champion C, Nabel EG, Luscher TF. Nitric oxide modulates expression of cell cycle regulatory proteins: a cytostatic strategy for inhibition of human vascular smooth muscle cell proliferation. Circulation. 2000;101:1982–1989. [PubMed]
44. Musgrove EA, Lee CS, Cornish AL, Swarbrick A, Sutherland RL. Antiprogestin inhibition of cell cycle progression in T-47D breast cancer cells is accompanied by induction of the cyclin-dependent kinase inhibitor p21. Mol Endocrinol. 1997;11:54–66. [PubMed]
45. Gansauge S, Nussler AK, Beger HG, Gansauge F. Nitric oxide-induced apoptosis in human pancreatic carcinoma cell lines is associated with a G1-arrest and an increase of the cyclin-dependent kinase inhibitor p21WAF1/CIP1. Cell Growth Differ. 1998;9:611–617. [PubMed]
46. Sarkar R, Gordon D, Stanley JC, Webb RC. Dual cell cycle-specific mechanisms mediate the antimitogenic effects of nitric oxide in vascular smooth muscle cells. J Hypertens. 1997;15:275–283. [PubMed]
47. Pervin S, Singh R, Chaudhuri G. Nitric oxide-induced cytostasis and cell cycle arrest of a human breast cancer cell line (MDA-MB-231): potential role of cyclin D1. Proc Natl Acad Sci USA. 2001;98:3583–3588. [PubMed]
48. Kelly MR, Geigerman CM, Loo G. Epigallocatechin gallate protects U937 cells against nitric oxide-induced cell cycle arrest and apoptosis. J Cell Biochem. 2001;81:647–658. [PubMed]
49. Villalobo A. Nitric oxide and cell proliferation. FEBS J. 2006;273:2329–2344. [PubMed]
50. Chazotte-Aubert L, Pluquet O, Hainaut P, Ohshima H. Nitric oxide prevents gamma-radiation-induced cell cycle arrest by impairing p53 function in MCF-7 cells. Biochem Biophys Res Commun. 2001;281:766–771. [PubMed]
51. Radi R, Peluffo G, Alvarez MN, Naviliat M, Cayota A. Unraveling peroxynitrite formation in biological systems. Free Radic Biol Med. 2001;30:463–488. [PubMed]
52. Dowell FJ, Martin W. Interaction between per-oxynitrite and L-cysteine: effects on rat aorta. Eur J Pharmacol. 1998;344:183–190. [PubMed]
53. Failli P, Palmieri L, D’Alfonso C, Giovannelli L, Generini S, Rosso AD, et al. Effect of N-acetyl-L-cysteine on peroxynitrite and superoxide anion production of lung alveolar macrophages in systemic sclerosis. Nitric Oxide. 2002;7:277–282. [PubMed]
54. Brewer JW, Diehl JA. PERK mediates cell cycle exit during the mammalian unfolded protein response. Proc Natl Acad Sci USA. 2000;97:12625–12630. [PubMed]
55. Cullinan SB, Zhang D, Hannink M, Arvisais E, Kaufman RJ, Diehl JA. Nrf2 is a direct PERK substrate and effector of PERK-dependent cell survival. Mol Cell Biol. 2003;23:7198–7209. [PMC free article] [PubMed]
56. Raven JF, Koromilas AE. PERK and PKR: old kinases learn new tricks. Cell Cycle. 2008;7:1146–1150. [PubMed]
57. Moretti L, Cha YI, Niermann KJ, Lu B. Switch between apoptosis and autophagy: radiation-induced endoplasmic reticulum stress? Cell Cycle. 2007;6:793–798. [PubMed]
58. Jiang HY, Wek RC. Phosphorylation of the {alpha}-Subunit of the Eukaryotic Initiation Factor-2 (eIF2{alpha}) Reduces Protein Synthesis and Enhances Apoptosis in Response to Proteasome Inhibition. J Biol Chem. 2005;280:14189–14202. [PubMed]
59. Hamanaka RB, Bennett BS, Cullinan SB, Diehl JA. PERK and GCN2 contribute to eIF2alpha phosphorylation and cell cycle arrest after activation of the unfolded protein response pathway. Mol Biol Cell. 2005;16:5493–5501. [PMC free article] [PubMed]
60. Tvegard T, Soltani H, Skjolberg HC, Krohn M, Nilssen EA, Kearsey SE, et al. A novel checkpoint mechanism regulating the G1/S transition. Genes Dev. 2007;21:649–654. [PubMed]
61. Menacho-Marquez M, Perez-Valle J, Arino J, Gadea J, Murguia JR. Gcn2p regulates a G1/S cell cycle checkpoint in response to DNA damage. Cell Cycle. 2007;6:2302–2305. [PubMed]
62. Baltzis D, Pluquet O, Papadakis AI, Kazemi S, Qu LK, Koromilas AE. The eIF2alpha kinases PERK and PKR activate glycogen synthase kinase 3 to promote the proteasomal degradation of p53. J Biol Chem. 2007;282:31675–31687. [PubMed]
63. Will K, Neben M, Schmidt-Rose T, Deppert W, Wittern KP, Bergemann J. P53-dependent UVB responsiveness of human keratinocytes can be altered by cultivation on cell cycle-arrested dermal fibroblasts. Photochem Photobiol. 2000;71:321–326. [PubMed]