Differential Activation of Mitogen-activated Protein Kinases by Nitric Oxide-related Species

Abstract

Many studies have identified nitric oxide (NO) and related chemical species (NOx) as having critical roles in neurotransmission, vasoregulation, and cellular signaling. Previous work in this laboratory has focused on elucidating the mechanism of NOx signaling in cells. We have demonstrated that NOx-induced activation of the guanine nucleotide-binding protein p21ras leads to nuclear translocation of the transcription factor NFκB. Here, we investigated whether intermediary signaling elements, namely the mitogen-activated protein (MAP) kinases, are involved in mediating NOx signaling. We found that NOx activates the extracellular signal-regulated kinase (ERK), p38, and c-Jun NH2-terminal kinase (JNK) subgroups of MAP kinases in human Jurkat T cells. JNK was found to be 100-fold more sensitive to NOx stimulation than p38 and ERK. In addition, the activation of JNK and p38 by NOx was more rapid than ERK activation. Depletion of intracellular glutathione augmented the NOx-induced increase in kinase activity. Furthermore, endogenous NO, generated from NO synthase, activated ERK, and NOx-induced MAP kinase activation was effectively blocked by the farnesyl transferase inhibitor α-hydroxyfarnesylphosphonic acid. These data support the hypothesis that critical signaling kinases, such as ERK, p38, and JNK, are activated by NO-related species and thus participate in NO signal transduction. These findings establish a role for multiple MAP kinase signaling pathways in the cellular response to NOx. Many studies have identified nitric oxide (NO) and related chemical species (NOx) as having critical roles in neurotransmission, vasoregulation, and cellular signaling. Previous work in this laboratory has focused on elucidating the mechanism of NOx signaling in cells. We have demonstrated that NOx-induced activation of the guanine nucleotide-binding protein p21ras leads to nuclear translocation of the transcription factor NFκB. Here, we investigated whether intermediary signaling elements, namely the mitogen-activated protein (MAP) kinases, are involved in mediating NOx signaling. We found that NOx activates the extracellular signal-regulated kinase (ERK), p38, and c-Jun NH2-terminal kinase (JNK) subgroups of MAP kinases in human Jurkat T cells. JNK was found to be 100-fold more sensitive to NOx stimulation than p38 and ERK. In addition, the activation of JNK and p38 by NOx was more rapid than ERK activation. Depletion of intracellular glutathione augmented the NOx-induced increase in kinase activity. Furthermore, endogenous NO, generated from NO synthase, activated ERK, and NOx-induced MAP kinase activation was effectively blocked by the farnesyl transferase inhibitor α-hydroxyfarnesylphosphonic acid. These data support the hypothesis that critical signaling kinases, such as ERK, p38, and JNK, are activated by NO-related species and thus participate in NO signal transduction. These findings establish a role for multiple MAP kinase signaling pathways in the cellular response to NOx. INTRODUCTIONOur previous work focused on identifying the signaling cascade responsible for the positive regulatory effects of nitric oxide (NO) 1The abbreviations used are: NOnitric oxideNOxnitric oxide-related speciesSNPsodium nitroprussideSNAPS-nitroso-N-acetylpenicillamineERKextracellular signal-regulated kinaseJNKc-Jun NH2-terminal kinaseMAPmitogen-activated proteinBSOL-buthionine-(SR)-sulfoximineGSTglutathione S-transferase. and related chemical species (NOx) on human peripheral blood mononuclear cells (1Lander H.M. Sehajpal P. Levine D.M. Novogrodsky A. J. Immunol. 1993; 150: 1509-1516PubMed Google Scholar, 2Lander H.M. Sehajpal P.K. Novogrodsky A. J. Immunol. 1993; 151: 7182-7187PubMed Google Scholar). NOx was found to activate human lymphocytes, as evidenced by increased glucose uptake, tumor necrosis factor α secretion, and nuclear translocation of the transcription factor NFκB. Further studies demonstrated that NOx directly activates p21ras, leading to downstream events such as the activation of NFκB (3Lander H.M. Ogiste J.S. Pearce S.F.A. Levi R. Novogrodsky A. J. Biol. Chem. 1995; 270: 7017-7020Abstract Full Text Full Text PDF PubMed Scopus (333) Google Scholar). We have also identified p21ras as a general signaling target of reactive free radicals and suggested that it may be a cellular sensor for redox stress (4Lander H.M. Ogiste J.S. Teng K.K. Novogrodsky A. J. Biol. Chem. 1995; 270: 21195-21198Abstract Full Text Full Text PDF PubMed Scopus (368) Google Scholar). In those studies, reactive oxygen species were shown to signal downstream of p21ras by activation of the extracellular signal-regulated kinases (ERKs). Here, we examined whether activation of low molecular weight G proteins such as p21ras, Rac1, and Cdc42 by NOx also leads to activation of one or more of the mitogen-activated protein (MAP) kinase family members.The three identified mammalian MAP kinase subgroups include ERK (5Cano E. Mahadevan L.C. Trends Biochem. Sci. 1995; 20: 117-122Abstract Full Text PDF PubMed Scopus (996) Google Scholar), c-Jun NH2-terminal kinase (JNK; Refs. 6Dérijard B. Hibi M. Wu I.-H. Barrett T. Su B. Deng T. Karin M. Davis R. Cell. 1994; 76: 1025-1037Abstract Full Text PDF PubMed Scopus (2949) Google Scholar, 7Davis R. Trends Biochem. Sci. 1994; 19: 470-473Abstract Full Text PDF PubMed Scopus (917) Google Scholar, 8Kyriakis J.M. Banerjee P. Nikolakaki E. Dai T. Rubie E.A. Ahmad M.F. Avruch J. Woodjett J.R. Nature. 1994; 369: 156-160Crossref PubMed Scopus (2408) Google Scholar), and the recently identified p38 MAP kinase (9Raingeaud J. Gupta S. Rogers J.S. Dickens M. Han J. Ulevitch R.J. Davis R.J. J. Biol. Chem. 1995; 270: 7420-7426Abstract Full Text Full Text PDF PubMed Scopus (2038) Google Scholar, 10Freshney N.W. Rawlinson L. Guedson F. Jones E. Cowley S. Hsuan J. Saklatvala J. Cell. 1994; 78: 1039-1049Abstract Full Text PDF PubMed Scopus (774) Google Scholar, 11Han J. Lee J.D. Bibbs L. Ulevitch R.J. Science. 1994; 265: 808-811Crossref PubMed Scopus (2401) Google Scholar, 12Lee J.C. Laydon J.T. McDonnell P.C. Gallagher T.F. Kumar S. Green D. McNulty D. Blumenthal M.J. Heys J.R. Landvatter S.W. Strickler J.E. McLaughlin M.M. Siemens I.R. Fisher S.M. Livi G.P. White J.R. Adams J.L. Young P.R. Nature. 1994; 372: 739-746Crossref PubMed Scopus (3121) Google Scholar, 13Rouse J. Cohen P. Trigon S. Morange M. Alonso-Llamazares A. Zamanillo D. Hunt T. Nebreda A.R. Cell. 1994; 78: 1027-1037Abstract Full Text PDF PubMed Scopus (1494) Google Scholar). The well studied ERK group is typically activated by growth factors via a p21ras-dependent signal transduction pathway (5Cano E. Mahadevan L.C. Trends Biochem. Sci. 1995; 20: 117-122Abstract Full Text PDF PubMed Scopus (996) Google Scholar, 9Raingeaud J. Gupta S. Rogers J.S. Dickens M. Han J. Ulevitch R.J. Davis R.J. J. Biol. Chem. 1995; 270: 7420-7426Abstract Full Text Full Text PDF PubMed Scopus (2038) Google Scholar, 14Cobb M.H. Goldsmith E.J. J. Biol. Chem. 1995; 270: 14843-14846Abstract Full Text Full Text PDF PubMed Scopus (1657) Google Scholar). JNK and p38 MAP kinases respond to proinflammatory cytokines and environmental stress, although it is not clear whether this occurs via common or parallel pathways (5Cano E. Mahadevan L.C. Trends Biochem. Sci. 1995; 20: 117-122Abstract Full Text PDF PubMed Scopus (996) Google Scholar). The signal transduction pathways leading to ERK, JNK, and p38 kinase activation are biochemically and functionally distinct (7Davis R. Trends Biochem. Sci. 1994; 19: 470-473Abstract Full Text PDF PubMed Scopus (917) Google Scholar, 9Raingeaud J. Gupta S. Rogers J.S. Dickens M. Han J. Ulevitch R.J. Davis R.J. J. Biol. Chem. 1995; 270: 7420-7426Abstract Full Text Full Text PDF PubMed Scopus (2038) Google Scholar). Nevertheless, some extracellular stimuli can activate all of the MAP kinases simultaneously (5Cano E. Mahadevan L.C. Trends Biochem. Sci. 1995; 20: 117-122Abstract Full Text PDF PubMed Scopus (996) Google Scholar). In the present study, we attempt to further clarify the NOx-signaling pathway downstream of low molecular weight G proteins such as p21ras by exploring the effects of NO-related chemical species on the ERK, p38, and JNK subgroups of MAP kinase.DISCUSSIONPrevious work in this laboratory demonstrated a requirement for functional p21ras in activation of NFκB by NOx (3Lander H.M. Ogiste J.S. Pearce S.F.A. Levi R. Novogrodsky A. J. Biol. Chem. 1995; 270: 7017-7020Abstract Full Text Full Text PDF PubMed Scopus (333) Google Scholar). We also found that reactive oxygen intermediates signaled through a similar mechanism and thus generalized our findings on signal transduction by NOx to include reactive oxygen species (4Lander H.M. Ogiste J.S. Teng K.K. Novogrodsky A. J. Biol. Chem. 1995; 270: 21195-21198Abstract Full Text Full Text PDF PubMed Scopus (368) Google Scholar). In those studies, we found that reactive oxygen signals through p21ras, MAP kinase, and NFκB. We concluded from these studies that p21ras is directly targeted by reactive free radicals and may be a cellular sensor for redox status (4Lander H.M. Ogiste J.S. Teng K.K. Novogrodsky A. J. Biol. Chem. 1995; 270: 21195-21198Abstract Full Text Full Text PDF PubMed Scopus (368) Google Scholar).ERK, JNK, and p38 play critical intermediary roles in mediating signal transduction from the membrane to the nucleus (5Cano E. Mahadevan L.C. Trends Biochem. Sci. 1995; 20: 117-122Abstract Full Text PDF PubMed Scopus (996) Google Scholar, 7Davis R. Trends Biochem. Sci. 1994; 19: 470-473Abstract Full Text PDF PubMed Scopus (917) Google Scholar). Therefore, we investigated whether these signaling enzymes were activated by NOx and the NOx-generating compounds SNP and SNAP in human T cells. The molecular distinction between the different MAP kinase pathways lies in the substrate specificities of the MAP kinase kinases. There are six MAP kinase kinases in mammalian cells. MEK1, MEK2, and MEK5 activate ERK-related MAP kinases (29Ahn N.G. Seger R. Krebs E.G. Curr. Opin. Cell Biol. 1992; 4: 992-999Crossref PubMed Scopus (228) Google Scholar, 30Zhou G. Bao Z.Q. Dixon J.E. J. Biol. Chem. 1995; 270: 12665-12669Abstract Full Text Full Text PDF PubMed Scopus (535) Google Scholar); MKK3 and MKK6 activate p38 (31Derijard B. Raingeaud J. Barrett T. Wu I.-H. Han J. Ulevitch R.J. Davis R.J. Science. 1995; 267: 682-685Crossref PubMed Scopus (1405) Google Scholar, 32Raingeaud J. Barrett T. Davis R.J. Mol. Cell. Biol. 1996; 16: 1247-1255Crossref PubMed Scopus (1139) Google Scholar); and MKK4 activates JNK and p38 (31Derijard B. Raingeaud J. Barrett T. Wu I.-H. Han J. Ulevitch R.J. Davis R.J. Science. 1995; 267: 682-685Crossref PubMed Scopus (1405) Google Scholar, 33Sanchez I. Hughes R.T. Mayer B.J. Yee K. Woodgett J.R. Avruch J. Kyriakis J.M. Zon L.I. Nature. 1994; 372: 794-798Crossref PubMed Scopus (913) Google Scholar, 34Lin A. Minden A. Martinetto H. Claret F.-X. Lange-Carter C. Mercurio F. Johnson G.L. Karin M. Science. 1995; 268: 286-290Crossref PubMed Scopus (706) Google Scholar). These MAP kinase kinases are themselves activated by receptor tyrosine kinases and other mechanisms. Our findings suggest that NO-related species and reactive oxygen intermediates activate the MAP kinases through upstream activation, most likely by activation of a low molecular weight G protein such as p21ras, Rac1, or Cdc42 (22Coso O.A. Chiariello M. Yu J.-C. Teramoto H. Crespo P. Xu N. Miki T. Gutkind J.S. Cell. 1995; 81: 1137-1146Abstract Full Text PDF PubMed Scopus (1559) Google Scholar, 23Minden A. Lin A. Claret F.X. Abo A. Karin M. Cell. 1995; 81: 1147-1157Abstract Full Text PDF PubMed Scopus (1444) Google Scholar, 24Bagrodia S. Derijard B. Davis R.J. Cerione R. J. Biol. Chem. 1995; 270: 27995-27998Abstract Full Text Full Text PDF PubMed Scopus (598) Google Scholar, 25Zhang S. Han J. Sells M.A. Chernoff J. Kraus U.G. Ulevitch R.J. Bokoch G.M. J. Biol. Chem. 1995; 270: 23934-23936Abstract Full Text Full Text PDF PubMed Scopus (651) Google Scholar, 26Olson M.F. Ashworth A. Hall A. Science. 1995; 269: 1270-1272Crossref PubMed Scopus (1055) Google Scholar). The time course of activation of ERK by NOx was slow compared with the rapid effects of NOx on JNK and p38. This differential activation suggests that not all signals go through p21ras and that other related G proteins may be involved.JNK activation by UV light has been shown to be potentiated by the activation of p21Ha-ras (6Dérijard B. Hibi M. Wu I.-H. Barrett T. Su B. Deng T. Karin M. Davis R. Cell. 1994; 76: 1025-1037Abstract Full Text PDF PubMed Scopus (2949) Google Scholar). This finding demonstrates that JNK signals via at least one p21ras-dependent pathway. Because the effects of UV irradiation often mimic those of oxidative stress, it was postulated that the UV-induced activation of JNK may be initiated by oxidative stress signaling through an unknown redox sensor. We have previously suggested p21ras as a redox sensor in the free radical-induced activation of NFκB (4Lander H.M. Ogiste J.S. Teng K.K. Novogrodsky A. J. Biol. Chem. 1995; 270: 21195-21198Abstract Full Text Full Text PDF PubMed Scopus (368) Google Scholar). The present study describes the activation of JNK by some of the same reactive species. It is possible that p21ras or other low molecular weight G proteins (such as Rac1 and CDC42) serve similar roles in JNK activation.NO is produced in many cell types (35Nathan C. FASEB J. 1992; 6: 3051-3064Crossref PubMed Scopus (4132) Google Scholar). The effect of NO on cells depends on its local concentration, the redox status of its immediate environment, and the susceptibility of target sites for modification (18Stamler J.S. Singel D.J. Loscalzo J. Science. 1992; 258: 1898-1902Crossref PubMed Scopus (2435) Google Scholar, 36Gross S.S. Wolin M.S. Annu. Rev. Physiol. 1995; 57: 737-769Crossref PubMed Scopus (820) Google Scholar). Reactive nitrogen and oxygen overproduction is often associated with the synthesis of destructive species, such as superoxide, peroxynitrite, and nitrosonium ion. When these reactive species overwhelm the cell's antioxidant defenses, the resulting redox stress can cause lipid peroxidation, membrane damage, DNA breakage, and enzyme inactivation (20Halliwell B. Gutteridge J.M.C. Free Radicals in Biology and Medicine. 2nd Ed. Oxford University Press, New York1989Google Scholar). Our study suggests that cellular glutathione levels regulate a low molecular weight G protein-dependent nuclear signaling pathway in which the MAP kinases play an important role. Thus, the ability of NOx to trigger some signaling events within the cell, such as ERK, JNK, and p38 kinase activation, as shown herein, will likely be revealed under conditions in which cellular glutathione levels are depleted. INTRODUCTIONOur previous work focused on identifying the signaling cascade responsible for the positive regulatory effects of nitric oxide (NO) 1The abbreviations used are: NOnitric oxideNOxnitric oxide-related speciesSNPsodium nitroprussideSNAPS-nitroso-N-acetylpenicillamineERKextracellular signal-regulated kinaseJNKc-Jun NH2-terminal kinaseMAPmitogen-activated proteinBSOL-buthionine-(SR)-sulfoximineGSTglutathione S-transferase. and related chemical species (NOx) on human peripheral blood mononuclear cells (1Lander H.M. Sehajpal P. Levine D.M. Novogrodsky A. J. Immunol. 1993; 150: 1509-1516PubMed Google Scholar, 2Lander H.M. Sehajpal P.K. Novogrodsky A. J. Immunol. 1993; 151: 7182-7187PubMed Google Scholar). NOx was found to activate human lymphocytes, as evidenced by increased glucose uptake, tumor necrosis factor α secretion, and nuclear translocation of the transcription factor NFκB. Further studies demonstrated that NOx directly activates p21ras, leading to downstream events such as the activation of NFκB (3Lander H.M. Ogiste J.S. Pearce S.F.A. Levi R. Novogrodsky A. J. Biol. Chem. 1995; 270: 7017-7020Abstract Full Text Full Text PDF PubMed Scopus (333) Google Scholar). We have also identified p21ras as a general signaling target of reactive free radicals and suggested that it may be a cellular sensor for redox stress (4Lander H.M. Ogiste J.S. Teng K.K. Novogrodsky A. J. Biol. Chem. 1995; 270: 21195-21198Abstract Full Text Full Text PDF PubMed Scopus (368) Google Scholar). In those studies, reactive oxygen species were shown to signal downstream of p21ras by activation of the extracellular signal-regulated kinases (ERKs). Here, we examined whether activation of low molecular weight G proteins such as p21ras, Rac1, and Cdc42 by NOx also leads to activation of one or more of the mitogen-activated protein (MAP) kinase family members.The three identified mammalian MAP kinase subgroups include ERK (5Cano E. Mahadevan L.C. Trends Biochem. Sci. 1995; 20: 117-122Abstract Full Text PDF PubMed Scopus (996) Google Scholar), c-Jun NH2-terminal kinase (JNK; Refs. 6Dérijard B. Hibi M. Wu I.-H. Barrett T. Su B. Deng T. Karin M. Davis R. Cell. 1994; 76: 1025-1037Abstract Full Text PDF PubMed Scopus (2949) Google Scholar, 7Davis R. Trends Biochem. Sci. 1994; 19: 470-473Abstract Full Text PDF PubMed Scopus (917) Google Scholar, 8Kyriakis J.M. Banerjee P. Nikolakaki E. Dai T. Rubie E.A. Ahmad M.F. Avruch J. Woodjett J.R. Nature. 1994; 369: 156-160Crossref PubMed Scopus (2408) Google Scholar), and the recently identified p38 MAP kinase (9Raingeaud J. Gupta S. Rogers J.S. Dickens M. Han J. Ulevitch R.J. Davis R.J. J. Biol. Chem. 1995; 270: 7420-7426Abstract Full Text Full Text PDF PubMed Scopus (2038) Google Scholar, 10Freshney N.W. Rawlinson L. Guedson F. Jones E. Cowley S. Hsuan J. Saklatvala J. Cell. 1994; 78: 1039-1049Abstract Full Text PDF PubMed Scopus (774) Google Scholar, 11Han J. Lee J.D. Bibbs L. Ulevitch R.J. Science. 1994; 265: 808-811Crossref PubMed Scopus (2401) Google Scholar, 12Lee J.C. Laydon J.T. McDonnell P.C. Gallagher T.F. Kumar S. Green D. McNulty D. Blumenthal M.J. Heys J.R. Landvatter S.W. Strickler J.E. McLaughlin M.M. Siemens I.R. Fisher S.M. Livi G.P. White J.R. Adams J.L. Young P.R. Nature. 1994; 372: 739-746Crossref PubMed Scopus (3121) Google Scholar, 13Rouse J. Cohen P. Trigon S. Morange M. Alonso-Llamazares A. Zamanillo D. Hunt T. Nebreda A.R. Cell. 1994; 78: 1027-1037Abstract Full Text PDF PubMed Scopus (1494) Google Scholar). The well studied ERK group is typically activated by growth factors via a p21ras-dependent signal transduction pathway (5Cano E. Mahadevan L.C. Trends Biochem. Sci. 1995; 20: 117-122Abstract Full Text PDF PubMed Scopus (996) Google Scholar, 9Raingeaud J. Gupta S. Rogers J.S. Dickens M. Han J. Ulevitch R.J. Davis R.J. J. Biol. Chem. 1995; 270: 7420-7426Abstract Full Text Full Text PDF PubMed Scopus (2038) Google Scholar, 14Cobb M.H. Goldsmith E.J. J. Biol. Chem. 1995; 270: 14843-14846Abstract Full Text Full Text PDF PubMed Scopus (1657) Google Scholar). JNK and p38 MAP kinases respond to proinflammatory cytokines and environmental stress, although it is not clear whether this occurs via common or parallel pathways (5Cano E. Mahadevan L.C. Trends Biochem. Sci. 1995; 20: 117-122Abstract Full Text PDF PubMed Scopus (996) Google Scholar). The signal transduction pathways leading to ERK, JNK, and p38 kinase activation are biochemically and functionally distinct (7Davis R. Trends Biochem. Sci. 1994; 19: 470-473Abstract Full Text PDF PubMed Scopus (917) Google Scholar, 9Raingeaud J. Gupta S. Rogers J.S. Dickens M. Han J. Ulevitch R.J. Davis R.J. J. Biol. Chem. 1995; 270: 7420-7426Abstract Full Text Full Text PDF PubMed Scopus (2038) Google Scholar). Nevertheless, some extracellular stimuli can activate all of the MAP kinases simultaneously (5Cano E. Mahadevan L.C. Trends Biochem. Sci. 1995; 20: 117-122Abstract Full Text PDF PubMed Scopus (996) Google Scholar). In the present study, we attempt to further clarify the NOx-signaling pathway downstream of low molecular weight G proteins such as p21ras by exploring the effects of NO-related chemical species on the ERK, p38, and JNK subgroups of MAP kinase.