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Papillary thyroid cancer (PTC) is the most common type of thyroid cancer. Deciphering the pathophysiological mechanisms that contribute to PTC development is essential to the discovery of optimal diagnostic and therapeutic approaches. MiR-146b-5p has been identified as a cancer-associated microRNA highly up-regulated in PTC. This study explores the hypothesis that miR-146b-5p contributes to papillary thyroid carcinogenesis through regulation of cell signaling pathways in a manner that overcomes the cellular growth suppressive events and provides survival advantage. The effect of miR-146b-5p inhibition on major cancer related signaling pathways and expression of Stanniocalcin-1 (STC1), an emerging molecule associated with stress response and carcinogenesis, was tested in cultured primary thyroid cells using luciferase reporter assays, quantitative real-time PCR, immunofluorescence staining, and flow cytometry. Our results demonstrated that miR-146b-5p inhibits the JNK/AP1 pathway activity and down-regulates the expression of STC-1 in thyroid-cultured cells and in thyroid tissue samples. In the presence of miR-146b-5p, PTC cells were resistant to cell death in response to oxidative stress. This is a novel report that miR-146b-5p directly targets STC1 and regulates the activity of JNK/AP1 pathway. Considering the importance of the JNK/AP1 pathway and STC1 in mediating many physiological and pathological processes like apoptosis, stress response and cellular metabolism, a biological regulator of these pathways would have a great scientific and clinical significance. [ABSTRACT FROM AUTHOR]

The cardiac fibroblast is a remarkably versatile cell type that coordinates inflammatory, fibrotic and hypertrophic responses in the heart through a complex array of intracellular and intercellular signaling mechanisms. One important signaling node that has been identified involves p38 MAPK; a family of kinases activated in response to stress and inflammatory stimuli that modulates multiple aspects of cardiac fibroblast function, including inflammatory responses, myofibroblast differentiation, extracellular matrix turnover and the paracrine induction of cardiomyocyte hypertrophy. This review explores the emerging importance of the p38 MAPK pathway in cardiac fibroblasts, describes the molecular mechanisms by which it regulates the expression of key genes, and highlights its potential as a therapeutic target for reducing adverse myocardial remodeling.

Objective To investigate the changes in levels of c-Jun N-terminal kinase /stress-activated protein kinase (JNK/SAPK)in the lung tissues of rats with chronic obstructive pulmonary disease (COPD).Methods Sixteen Ⅱ male Wistar rats were randomly divided into two groups:the control group and COPD group. The control group was not treated, while the COPD group was stimulated with smoke to set up COPD models. After models were established, we detected the lung pathology through optical microscope, pulmonary function through animal pulmonary instrument, TNF-αexpression in lung tissue through ELISA, JNK/SAPK expression through immunohistochemistry, and JNK mRNA expression through RT-PCR.Results In the COPD group, light microscope showed that there was a large number of inflammatory cell infiltra-tion, alveolar fusion, and alveolar reduction, which proved the COPD model was successful. Compared with the control group, FEV 0.3 /FVC decreased, Ri and Re increased, and Cldyn decreased (P <0.05 or P <0.01),and TNF-αin-creased in the COPD group (P <0.05).Immunohistochemistry showed that JNK/SAPK expression improved in the COPD group. RT-PCR result showed JNK mRNA expression increased in the COPD group as compared with that of the control group (all P <0.05).JNK mRNA expression was positively correlated with TNF-α(r =0.751, P <0.05).Conclusion The level of JNK/SAPK increases in smoke-stimulated COPD rats, which can cause airway inflammation and emphysema, and promote the occurrence of COPD through activating smoke-induced inflammatory cytokines. [ABSTRACT FROM AUTHOR]

The sensing of stress signals and their transduction into appropriate responses are crucial for the adaptation, survival and infection of phytopathogenic fungi and oomycetes. Amongst evolutionarily conserved pathways, mitogen-activated protein kinase ( MAPK) cascades function as key signal transducers that use phosphorylation to convey information. In this study, we identified a gene, designated PsMPK7, one of 14 predicted genes encoding MAPKs in Phytophthora sojae. PsMPK7 was highly transcribed in each tested stage, but was up-regulated in the zoospore, cyst and cyst germination stages. Silencing of PsMPK7 affected the growth of germinated cysts, oospore production and the pathogenicity of soybean. PsMPK7 transcription was induced by stresses from sorbitol, NaCl and hydrogen peroxide. Transformants in which PsMPK7 expression was silenced ( PsMPK7-silenced) were significantly more sensitive to osmotic and oxidative stress. Aniline blue and diaminobenzidine staining revealed that the silenced lines did not suppress the host reactive oxygen species ( ROS) burst, indicating that either the inoculated plants activated stronger defence responses to the transformants and/or the PsMPK7-silenced transformants failed to overcome plant defences. In addition, extracellular secretion of laccase decreased in the silenced lines. Overall, our results indicate that the PsMPK7 gene encodes a stress-associated MAPK in P. sojae that is important not only for responses to various stresses, but also for ROS detoxification, cyst germination, sexual oospore production and infection of soybean. [ABSTRACT FROM AUTHOR]

Background: Advanced hepatocellular carcinoma (HCC) responds poorly to conventional systemic therapies. Therefore, new effective therapy strategies are urgently needed. Molecular targeted therapies have entered the field of anti-neoplastic treatment and are being used on their own and in combination with other drugs. Sorafenib inhibits proliferation and angiogenesis of HCC by suppressing the Raf serine/threonine kinases and the receptor tyrosine kinases. The proteasome inhibitor bortezomib has shown activity in a variety of solid tumors, including HCC. However, the precise anti-proliferative mechanisms of these agents remain unclear. Methods: We treated human hepatoma cell lines (Huh7 and Hep3B) and immortalized human hepatocyte (OUMS29) with sorafenib and/or proteasome inhibitors, including epoxomicin and acetyl-leucyl-leucyl-norleucinal. Cytotoxic effects were examined by morphometric analyses of apoptosis and necrosis. Apoptosis was also evaluated by Western blotting of keratin18, PARP and caspase3. The activity of Akt and stress-activated protein kinases was examined by Western blotting. Results: Both sorafenib and proteasome inhibitors induced apoptosis in Huh7 and OUMS29. However, sorafenib attenuated proteasome inhibitor-induced apoptosis. Sorafenib induced necrosis, especially in combination with proteasome inhibitors. Sorafenib induced down-regulation of Akt synergistically in combination with proteasome inhibitors in Huh7. Sorafenib inhibited both the JNK and p38 pathways in a time- and dose-dependent manner. In addition, sorafenib also inhibited proteasome inhibitor-mediated JNK and p38 activation in both Huh7 and OUMS29. Conclusions: Sorafenib enhances the anti-proliferative effect of proteasome inhibitors in part by inactivating the Akt signaling pathway and modulating stress-activated protein kinases. The combination of these agents could be an ideal molecular targeted therapy for HCC. [ABSTRACT FROM AUTHOR]

In a "two-component system," extracellular stimuli are transmitted by the transfer of a phosphoryl group from a sensor histidine kinase to a response regulator (RR), a mechanism referred to as phosphorelay. In the fission yeast Schizosaccharomyces pombe, peroxide stress signals are transmitted by phosphorelay to the Mcs4 RR, which activates the Spc1 MAP kinase (MAPK) cascade. We previously demonstrated that a glycolytic enzyme, glyceraldehyde-3-phosphate dehydrogenase (GAPDH) physically interacts with Mcs4 and promotes phosphorelay signaling to Mcs4. Independently of the phosphorelay mechanism, Mcs4 also plays a critical role in osmostress signaling, as a part of the stable ternary complex with the Wis4 and Win1 MAPK kinase kinases (MAPKKKs). Interestingly, GAPDH dissociates from Mcs4 upon osmostress, while oxidative stress promotes their association. The Mcs4 RR may serve as a switching hub that mediates activation of the Wis4-Win1 MAPKKK heteromer in response to different forms of stress. [ABSTRACT FROM AUTHOR]

Here we investigated the effects of mineralocorticoid in the regulation of catecholamine biosynthesis using rat pheochromocytoma PC 12 cells. Expression of mineralocorticoid receptor (MR) was confirmed in undifferentiated PC 12 cells. Aldosterone stimulated dopamine production by PC12 cells without any increase in cAMP activity. Aldosterone-induced dopamine accumulation was enhanced in accordance with the increase in the rate-limiting enzyme tyrosine hydroxylase (TH). Blocking MR with eplerenone suppressed aldosterone-induced increases of TH mRNA and dopamine production. A glucocorticoid receptor (GR) antagonist, RU-486, attenuated dexamethasone- but not aldosterone-induced TH expression. Cycloheximide reduced both aldosterone- and dexamethasone-induced TH mRNA. A SAPK/JNK inhibitor, SP600 125, suppressed aldosterone-induced TH mRNA expression; however, the aldosterone-induced TH expression was not affected by inhibition of ERK½, p38-MAPK, Rho-kinase, PI 3-kinase, and PKC. It was of note that cotreatment with eplerenone and SP600125 restored aldosterone-induced TH mRNA expression to basal levels. To investigate the involvement of bone morphogenetic protein (BMP) actions in aldosterone-induced catecholamine production, we examined the effects of BMP-4 and BMP-7, which are expressed in the adrenal medulla, on catecholamine biosynthesis. BMP-4 preferentially enhanced aldosterone-induced TH mRNA and dopamine production, although BMP-4 alone did not affect TH expression. The BMP-4 enhancement of aldosterone-induced TH expression was not observed in cells treated with eplerenone. BMP-4 did not affect MR expression of PC12 cells; however, it did enhance aldosterone-induced SAPK/JNK phosphoiylation. Inhibition of SAPK/JNK or Rho suppressed BMP-4 enhancement of aldosterone-induced TH expression. Collectively, our findings demonstrate that aldosterone stimulates catecholamine biosynthesis in adrenomedullar cells via MR through genomic action and partly through nongenomic action by Rho-SAPK/JNK signaling, the latter of which is facilitated by BMP-4. A functional link between MR actions and endogenous BMP may be involved in the catecholamine production. [ABSTRACT FROM AUTHOR]

Abstract: Heavy metals are important environmental pollutants that affect many cellular functions. The signaling pathways that lead to apoptosis of adrenocortical cells following exposure to cadmium chloride (CdCl2) have not yet been fully clarified. We developed a primary culture of guinea pig adrenocortical cells and treated it with various concentrations of CdCl2 for different time periods. The apoptosis induced by CdCl2 was detected by fluorescein isothiocyanate-labeled annexin V and propidium iodide staining using a flow cytometer. Stress-activated protein kinase (SAPK) activities were measured by immunoprecipitation and chemiluminescence assay. After 2h of treatment, the apoptotic cell rate increased in a dose-dependent manner. The difference between the high-dose group and the control group was significant (P<0.01). The apoptosis was also found to increase in a time-dependent manner in cells treated with 50μmol/L CdCl2. Among the various treatment period groups, the difference between more than 1h-treated groups and the control group was significant (P<0.01). The SAPK activity increased with an increase in CdCl2 dose after 5min of exposure. However, after 15min of exposure to CdCl2, the SAPK activity decreased with an increase in exposure time. Our findings suggest that the SAPK signaling pathway might play an important role in CdCl2-induced apoptosis of adrenocortical cells. [Copyright &y& Elsevier]

We have shown previously that N-(4-hydroxyphenyl)retinamide (4HPR, fenretinide), a retinoic acid derivative, induces neuronal differentiation in cultured human retinal pigment epithelial (RPE) cells [Chen et al., J. Neurochem., 84 (2003), 972]. We asked the question whether the mitogen-activated protein kinase (MAPK) pathway is involved in the regulation of the 4HPR-induced neuronal differentiation of RPE (ARPE-19) cells. When we treated ARPE-19 cells with 4HPR, c-Raf and MEK1/2 kinase were activated resulting in activation of the downstream effector ERK1/2 and of SAPK/JNK. By blocking the upstream kinase MEK1/2 with specific inhibitor U0126 we abrogated the 4HPR-induced phosphorylation of ERK1/2 and SAPK/JNK, indicating that the neuronal differentiation occurs through a positive cross-talk between the ERK and the SAPK/JNK pathways. Both U0126 and the suppression of ERK1/2 expression with small interfering RNA effectively blocked the 4HPR-induced neuronal differentiation of RPE cells and the expression of calretinin. The activated ERK1/2 then induced a sequential activation of p90RSK, and increase in phosphorylation of transcription factors c- fos and c- jun leading to transcriptional activation of AP-1. Taken together, our results clearly demonstrate that c-Raf/MEK1/2 signaling cascade involving ERK1/2 plays a central role in mediating the 4HPR-induced neuronal differentiation and calretinin expression in the human ARPE-19 retinal pigment epithelial cell line. [ABSTRACT FROM AUTHOR]

Abstract: MAPK Pmk1p is the central element of a cascade involved in the maintenance of cell integrity and other functions in Schizosaccharomyces pombe. Pmk1p becomes activated by multiple stressing situations and also during cell separation. GTPase Rho2p acts upstream of the protein kinase C homolog Pck2p to activate the Pmk1 signalling pathway through direct interaction with MAPKKK Mkh1p. In this work we analyzed the functional significance of both Rho2p and Pck2p in the transduction of various stress signals by the cell integrity pathway. The results indicate that basal Pmk1p activity can be positively regulated by alternative mechanisms which are independent on the control by Rho2p and/or Pck2p. Unexpectedly, Pck1p, another protein kinase C homolog, negatively modulates Pmk1p basal activity by an unknown mechanism. Moreover, different elements appear to regulate the stress-induced activation of Pmk1p depending on the nature of the triggering stimuli. Whereas Pmk1p activation induced by hyper- or hypotonic stresses is channeled through Rho2p–Pck2p, other stressors, like glucose deprivation or cell wall disturbance, are transduced via other pathways in addition to that of Rho2p–Pck2p. On the contrary, Pmk1p activation observed during cell separation or after treatment with hydrogen peroxide does not involve Rho2p–Pck2p. Finally, Pck2p function is critical to maintain a Pmk1p basal activity that allows Pmk1p activation induced by heat stress. These data demonstrate the existence of a complex signalling network modulating Pmk1p activation in response to a variety of stresses in fission yeast. [Copyright &y& Elsevier]

Abstract: The cellular response to insult occurs by signaling via the stress-activated protein kinases, p38, and c-Jun NH2-terminal kinase (JNK). In the present study, we determined if hyperosmotic stress to rat hippocampal slices activates p38 and JNK and whether hyperosmolarity is a potential apoptotic stimulus in this experimental paradigm. Hyperosmotic stress, produced by addition of sorbitol to the incubation buffer, increased p38 phosphorylation; in contrast, JNK phosphorylation was not increased above control levels. Activation of p38 by phosphorylation was detected within 15 min of osmotic stress and was maintained through 360 min of hyperosmolarity. Concurrently, levels of cytochrome c were increased in the cytosolic fraction, indicating a decline in mitochondrial integrity. To a greater extent than the apoptotic stimulus, staurosporine, hyperosmotic stress activated caspase-3. Exposure to sorbitol also resulted in cleavage of the nuclear repair enzyme, poly(ADP-ribose) polymerase (PARP) and induced DNA fragmentation in slices. Concomitant treatment with sorbitol and SB202190, a selective p38 inhibitor, prevented the increase in cytosolic cytochrome c, decreased caspase-3 activation, and partially reduced PARP cleavage in a concentration-dependent manner. Inhibition of JNK with SP600125 did not affect caspase-3 activation in hippocampal slices. These results reveal hyperosmotic stress to be a potent activator of caspase-3 in hippocampus and suggest that downstream correlates of p38 signaling through a mitochondrial apoptotic pathway contribute significantly in the response to this insult. [Copyright &y& Elsevier]

Abstract: The environmental neurotoxin arsenic has recently been associated with altered neurofilament (NF) content in sciatic nerve. We examined herein the impact of sodium arsenite (the inorganic form of arsenic) on NF dynamics. Treatment of differentiated NB2/d1 cells and cultured dorsal root ganglion neurons decreased NF transport into axonal neurites and increased perikaryal phospho-NF immunoreactivity. Both of these effects were prevented by a pharmacological inhibitor (SP600125) of c-jun terminal kinase and by expression of a dominant-negative form of this kinase. Arsenic-induced inhibition of NF transport was prevented by treatment with lithium, a selective inhibitor of glycogen synthase kinase-3beta. Pharmacological inhibitors of cyclin-dependent kinase 5 and p38 mitogen-activated protein kinase did not attenuate the effects of arsenic on NF dynamics. These latter findings suggest that this environmental neurotoxin could contribute to peripheral neuropathy by perturbing NF dynamics. [Copyright &y& Elsevier]

It has now been established through multiple lines of evidence that oxidative stress is an early event in Alzheimer's disease, occurring prior to the canonical cytopathology. Thus, oxidative stress likely plays a key pathogenic role in the disease and is clearly involved in the cell loss and other neuropathology associated with Alzheimer's disease as demonstrated by the large number of metabolic signs of oxidative stress and by markers of oxidative damage. One puzzling observation, however, is that oxidative damage decreases with disease progression, such that levels of markers of rapidly formed oxidative damage, which are initially elevated, decrease as the disease progresses to advanced Alzheimer's disease. This finding indicates that reactive oxygen species not only cause damage to cellular structures but also provoke cellular responses, such as the compensatory upregulation of antioxidant enzymes found in vulnerable neurons in Alzheimer's disease. Not surprisingly, stress-activated protein kinase pathways, which are activated by oxidative stress, are extensively activated during Alzheimer's disease. In this review, we present the evidence of oxidative stress and compensatory responses that occur in Alzheimer's disease with a particular focus on the roles and mechanism of activation of stress-activated protein kinase pathways. [ABSTRACT FROM AUTHOR]

Abstract: c-Jun N-terminal kinase (JNK), along with its upstream activator MEKK-1, is typically thought of as a stress-activated kinase that mediates apoptosis. However, additional studies indicate that the MEKK-1/JNK pathway mediates critical aspects of neuronal survival and differentiation. Herein, we demonstrate that transfection of differentiated NB2a/d1 cells with a construct expression constitutively activated (ca) MEKK-1 increases levels of phospho-dependent neurofilament (NF) immunoreactivity within perikarya, while expression of a dominant-negative (dn) form of MEKK-1 decreases it. Steady-state levels of perikaryal phospho-NF immunoreactivity are reduced and the increase resulting from expression of caMEKK-1 is prevented, by the JNK inhibitor SP600125, suggesting that JNK is a major downstream effector of MEKK-1 on NF phosphorylation. Unexpectedly, both caMEKK-1 and dnMEKK-1 inhibited neuritogenesis as well as translocation of NFs into newly elaborated neurites. The JNK inhibitor SP600125 also inhibited NF transport in a dose-dependent manner. caMEKK-1 also prevented the increase in NF transport otherwise mediated by MAP kinase. Finally, both caMEKK-1 and dnMEKK-1 prevented initial neuritogenesis. These findings indicate that the MEKK-1/JNK pathway regulates critical aspects of initial outgrowth, and subsequent stabilization of axonal neurites. [Copyright &y& Elsevier]

Abstract: Mitogen-activated protein (MAP) kinases are a family of serine/threonine protein kinases that play an important role in a myriad of cellular processes, including cell proliferation, differentiation, and apoptosis. Abnormal activation of MAP kinases has been shown to participate in a variety of human diseases which include cancer, septic shock, rheumatoid arthritis, diabetes, and cardiovascular diseases. Active MAP kinase enzymes are not only valuable for basic biomedical research but are also critical for the development of pharmacological inhibitors as therapeutic drugs in the treatment of relevant human diseases. MAP kinases produced in a bacterial system are poorly active due to a lack of proper phosphorylation at their characteristic threonine and tyrosine residues. To overcome these limitations, we have developed a mammalian expression system for high level expression and one-step purification of enzymatically MAP kinases. We cloned JNK1, p38, and p38-regulated MAP kinase-activated protein kinase-2 into the mammalian expression vector pEBG, and expressed these protein kinases as glutathione S-transferase fusion proteins in human embryonic kidney 293T cells through transient transfection. The protein kinases were activated in vivo through treating the transfected cells with sodium arsenite and affinity-purified using glutathione–Sepharose beads. The enzymatic activities of these protein kinases were demonstrated by Western blot analysis and in vitro kinase assays. Our results indicate that this system is an extremely powerful tool for generating valuable reagents, and could be very valuable for proteomic studies. [Copyright &y& Elsevier]

Abstract: In Alzheimer''s disease, the complex catabolism of amyloid precursor protein (APP) leads to the production of amyloid-beta (Abeta) peptide, the major component of amyloid deposits. APP is cleaved by beta- and alpha-secretases to generate APP carboxy-terminal fragments (CTFs). Abeta peptide and amyloid intracellular domain are resulting from the cleavage of APP-CTFs by the gamma-secretase. In the present study, we hypothesize that post-translational modification of APP-CTFs could modulate their processing by the gamma-secretase. Inhibition of the gamma-secretase was shown to increase the total amount of APP-CTFs. Moreover, we showed that this increase was more marked among the phosphorylated variants and directly related to the activity of the gamma-secretase, as shown by kinetics analyses. Phosphorylated CTFs were shown to associate to presenilin 1, a major protein of the gamma-secretase complex. The phosphorylation of CTFs at the threonine 668 resulting of the c-Jun N-terminal kinase activation was shown to enhance their degradation by the gamma-secretase. Altogether, our results demonstrated that phosphorylated CTFs can be the substrates of the gamma-secretase and that an increase in the phosphorylation of APP-CTFs facilitates their processing by gamma-secretase. [Copyright &y& Elsevier]

Abstract: c-Jun N-terminal kinase (JNK) activation is linked to the aberrant cell death in several neurodegenerative disorders, including Parkinson’s and Alzheimer’s disease. The sequence similarity among the JNK isoforms and fellow MAP kinase family member p38 has rendered the challenge of producing JNK3-specific inhibitors difficult. Using the crystal structure of JNK3 complexed with JNK inhibitors, potential compound-interacting amino acid residues were mutated to the corresponding residues in p38. The effects of these mutations on the kinetic parameters with three compounds were examined: a JNK3- (vs. p38-) selective inhibitor (SP 600125); a p38-selective inhibitor (Merck Z); and a potent combined JNK3 and p38 inhibitor (Merck Y). The data confirm the role of the JNK3 residues Ile-70 and Val-196 in both inhibitor and ATP-binding. Remarkably, the Ile-70-Val and Val-196-Ala mutations caused an increase and decrease, respectively, in the binding affinity of the p38-specific compound, Merck Z, of 10-fold. The Ile-70-Val effect may be due to the increased capacity of the active site to accommodate Merck Z, whereas the Val-196-Ala mutant may induce an unfavourable conformational change. Conservative mutations of the Asn-152 and Gln-155 residues inactivated the JNK3 enzyme, possibly interfering with protein folding in a critical hinge region of the protein. [Copyright &y& Elsevier]

The c-Jun NH2-terminal kinase isoform (JNK) 1 is implicated in type 2 diabetes However, a potential role for the JNK2 protein kinase in diabetes has not been established. Here, we demonstrate that JNK2 may play an important role in type 1 (insulin-dependent) diabetes that is caused by autoimmune destruction of β cells. Studies of nonobese diabetic mice demonstrated that disruption of the Mapk9 gene (which encodes the JNK2 protein kinase) decreased destructive insulitis and reduced disease progression to diabetes. CD4+ T cells from JNK2-deficient nonobese diabetic mice produced less IFN-&#x03B3 but significantly increased amounts of IL-4 and IL-5. indicating polarization toward the TH2 phenotype. This role of JNK2 to control the Thl/Th2 balance of the immune response represents a mechanism of protection against autoimmune diabetes. We conclude that JNK protein kinases may have important roles in diabetes, including functions of JNK1 in type 2 diabetes and JNK2 in type 1 diabetes. [ABSTRACT FROM AUTHOR]

Acute pancreatitis is associated with stress kinase activation and cytokine production. We hypothesize that bile-pancreatic juice exclusion activates p38MAPK and induces TNF-α production in ligation-induced acute pancreatitis. We compared rats with 1–3 h of duct ligation, duct ligation with duodenal bile-pancreatic juice replacement from a donor rat, and sham operation. Pancreatic homogenates were analyzed as follows: (a) Immunoblots using phospho-specific p38MAPK antibody showed increased p38MAPK activation after ligation that was inhibited by bile-pancreatic juice replacement. (b) Immune-complex kinase assay using ATF-2 as substrate showed increased p38MAPK activation after ligation that was subdued by bile-pancreatic juice replacement. (c) ELISA showed increased pancreatic TNF-α production after ligation that was significantly ameliorated by bile-pancreatic juice replacement. Conclusion: Bile-pancreatic juice exclusion from gut increases p38MAPK activation and TNF-α production in this experimental model. Our findings support our central hypothesis that bile-pancreatic juice exclusion exacerbates cell stress and acute inflammation in ligation-induced acute pancreatitis. Copyright © 2005 S. Karger AG, Basel and IAP [ABSTRACT FROM AUTHOR]

Microtubule-associated protein tau in a hyperphosphorylated state is the major component of the filamentous lesions that define a number of neurodegenerative diseases commonly referred to as tauopathies. Hyperphosphorylation of tau at most sites appears to precede filament assembly. Many of the hyperphosphorylated sites are serine/threonine–proline sequences. Here we show that c-Jun N-terminal kinases JNK1, JNK2 and JNK3 phosphorylate tau at many serine/threonine–prolines, as assessed by the generation of the epitopes of phosphorylation-dependent anti-tau antibodies. Of the three protein kinases, JNK2 phosphorylated the most sites in tau, followed by JNK3 and JNK1. Phosphorylation by JNK isoforms resulted in a greatly reduced ability of tau to promote microtubule assembly. These findings extend the number of candidate protein kinases for the hyperphosphorylation of tau in Alzheimer's disease and other neurodegenerative disorders. [ABSTRACT FROM AUTHOR]

Previously, we have shown that ceramide is able to directly bind to and activate c-Raf and to trigger the downstream classical mitogen-activated protein kinase (MAPK/ERK) cascade in glomerular mesangial cells [Proc. Natl. Acad. Sci. USA 93 (1996) 6959]. In this study, we show that ceramide acts differently in glomerular endothelial cells in that treatment of endothelial cells with exogenous ceramide leads to a potent activation of the stress-activated protein kinase (SAPK/JNK) cascade but not to an activation of the classical ERK cascade. A similar effect was observed with the inflammatory cytokines TNFαand IL-1β,which activate a sphingomyelinase and thereby increase intracellular ceramide levels. The activation of JNKs as shown by c-Jun phosphorylation assays was paralleled by increased phosphorylation of the two JNK isoforms, p45 and p54. In addition, also the activator of JNKs, SEK1, was found to be increasingly phosphorylated by exogenous ceramide as well as by TNFα. In contrast, dihydroceramide had no effect on JNK or SEK1 phosphorylation. To see whether ceramide directly binds to MEKK1, which is the c-Raf analog in the SAPK cascade, a radioiodinated photoaffinity labeling analogue of ceramide, (N-[3-[[[2-(125I)iodo-4-[3-(trifluoromethyl)-3H-diazirin-3-yl]benzyl]oxy]-carbonyl] propanoyl]-d-erythro-sphingosine) ([125I]TID-ceramide) was used. Stimulation of endothelial cells with this [125I]TID-ceramide for 5 min followed by a short photolysis defined MEKK1 as a direct target of ceramide. With the same method, protein kinase C-α (PKC-α) was identified as a ceramide target. In contrast, no binding to c-Raf or the MEKK1 activator p65-PAK could be detected. A direct binding of ceramide to MEKK1 was also confirmed by affinity chromatography using a ceramide-coupled sepharose column. Furthermore, the ceramide-activated SAPK/JNK cascade is clearly involved in the mechanism of apoptosis, since in the presence of a JNK inhibitor, ceramide-induced DNA fragmentation is significantly reduced.In summary, we have shown that ceramide potently activates the SAPK cascade but not the ERK cascade in endothelial cells, which contrasts to mesangial cells where ceramide activates the ERK pathway and has only a minor effect on the SAPK cascade. Regarding the direct target of ceramide binding and action in endothelial cells, we identified MEKK1 as a further member of the growing family of ceramide-activated protein kinases. [Copyright &y& Elsevier]

Multiple lines of evidence demonstrate that oxidative stress is an early event in Alzheimer''s disease (AD), occurring prior to cytopathology, and therefore may play a key pathogenic role in the disease. Indeed, that oxidative mechanisms are involved in the cell loss and other neuropathology associated with AD is evidenced by the large number of metabolic signs of oxidative stress as well as by markers of oxidative damage. However, what is intriguing is that oxidative damage decreases with disease progression, such that levels of markers of rapidly formed oxidative damage, which are initially elevated, decrease as the disease progresses to advanced AD. This finding, along with the compensatory upregulation of antioxidant enzymes found in vulnerable neurons in AD, indicates that reactive oxygen species (ROS) not only cause damage to cellular structures but also provoke cellular responses. Mammalian cells respond to extracellular stimuli by transmitting intracellular instructions by signal transduction cascades to coordinate appropriate responses. Therefore, not surprisingly stress-activated protein kinase (SAPK) pathways, pathways that are activated by oxidative stress, are extensively activated during AD. In this paper, we review the evidence of oxidative stress and compensatory responses that occur in AD with a particular focus on the roles and mechanism of activation of SAPK pathways. [Copyright &y& Elsevier]

The adenosine A3 receptor generally couples to the Gi class of heterotrimeric G proteins, thereby decreasing cAMP levels and also mediating signaling via release of βγ subunits. Here we describe the central role of phosphatidylinositol-3′-kinase (PI3K) for adenosine A3 receptor-induced intracellular signaling to the stress-activated protein kinase p38 and the extracellular signal-regulated protein kinases ERK1/2. We used Chinese hamster ovary cells expressing the human adenosine A3 receptor, phospho-specific antibodies and different pharmacological tools to dissect the signaling pathways involving PI3K. The adenosine receptor agonist 5′N-ethylcarboxamidoadenosine induced a time- and dose-dependent increase in p38 and ERK1/2 phosphorylation, two signaling pathways that appeared also to be activated in the immortalized microglia cell line N13, which expressed endogenous adenosine A3 receptors. The 5′N-ethylcarboxamidoadenosine-induced effects on p38 and ERK1/2 in CHO cells were blocked by pertussis toxin pretreatment and were sensitive to pharmacological inhibition of PI3K. In addition, inhibition of Rac/Cdc42, small GTPases of the Rho family, by clostridium toxin B, diminished p38 phosphorylation but did not affect ERK1/2. Furthermore, we identified the serine 727 site of signal transducer and activator of transcription STAT3 as a probable downstream target of ERK1/2, and thereby provide evidence that adenosine A3 receptor mediated ERK1/2 activation has functional consequences. [Copyright &y& Elsevier]

: BackgroundCholecystokinin-A (CCK-A) and cholinergic receptor pathways, capable of activating stress kinases p38 mitogen-activated protein kinase (p38MAPK) and cJUN N-terminal kinase (JNK), are implicated in the pathogenesis of ligation-induced acute pancreatitis in rats. As ligation-induced acute pancreatitis in rats is associated with CCK-A receptor induction and p38MAPK activation, and as receptor induction could amplify acinar hyperstimulation and exacerbate cell stress, we tested the hypothesis that the cholinergic M3 receptor is induced and JNK is activated in this model.: MethodsCholinergic M3 receptor expression and JNK activation was compared in rats 1, 3, or 24 hours after sham operation or duct ligation.: ResultsImmunoblot analysis of pancreatic homogenates showed a time-dependent increase in cholinergic M3 receptor protein, total JNK, and phospho-JNK after duct ligation.: ConclusionsThere is a rapid and progressive cholinergic M3 receptor induction and JNK activation in ligation-induced acute pancreatitis in rats. These findings may have significance in the mechanism of disease pathogenesis. [Copyright &y& Elsevier]

The present study examined the existence of the adenosine A1, A2A, and A2B receptors and the effect of receptor activation on cAMP accumulation and protein phosphorylation in primary rat skeletal muscle cells. Presence of mRNA and protein for all three receptors was demonstrated in both cultured and adult rat skeletal muscle. NECA (10−9–10−4 M) increased the cAMP concentration in cultured muscle cells with an EC50 of (95% confidence interval)=15 (5.9–25.1) μM, whereas CGS 21680 (10−9–10−4 M) had no effect on cAMP accumulation. Concentrations of [R]-PIA below 10−6 M had no effect on cAMP accumulation induced by either isoproterenol or forskolin. NECA resulted in phosphorylation of CREB with an EC50 of (95% confidence interval)=1.7 (0.40–7.02) μM, whereas ERK1/2 and p38 phosphorylation was unchanged. The results show that, although the A1, A2A, and A2B receptors are all present in skeletal muscle cells, the effect of adenosine on adenylyl cyclase activation and phosphorylation of CREB is mainly mediated via the adenosine A2B receptor. [Copyright &y& Elsevier]

MAPK/ERK kinase kinase 2 (MEKK2) is a member of the mitogen-activated protein kinase kinase kinase (MAP3K) family of protein kinases. MAP3Ks are components of a three-tiered protein kinase pathway in which a MAP3K phosphorylates and activates a mitogen-activated protein kinase kinase (MAP2K), which in turn activates a mitogen-activated protein kinase (MAPK). We have previously identified residues within protein kinase subdomain X in the MAP3K, MEKK1, that are critical for its interaction with the MAP2K, MKK4, and MEKK1-induced MKK4 activation. We report here that kinase subdomain X also plays a critical role in MEKK2 activity. Select point mutations in subdomain X impair MEKK2 phosphorylation of the MAP2Ks, MKK7 and MEK5, abolish MEKK2-induced activation of the MAPKs, JNK1 and ERK5, and diminish MEKK2-dependent activation of an AP-1 reporter gene. Interestingly, the spectrum of mutations in subdomain X of MEKK2 that affects its activity is overlapping with but not identical to those that have effects on MEKK1. Thus, mutations in subdomain X differentially affect MEKK2 and MEKK1. [Copyright &y& Elsevier]

Bcl10 is a signaling protein required for activation of the NF-κB transcription factor. NF-κB is an important mediator of genotoxic stress and regulates the expression of genes required for both cell proliferation and cell death. Bcl10 is phosphorylated in vivo, however, the protein kinase or kinases responsible are not known. Here, we show that Bcl10 is phosphorylated in response to UV irradiation. UV-induced phosphorylation of Bcl10 was inhibited by the p38 stress-activated protein kinase inhibitors SB203580 and PD169316, suggesting that p38 is required for UV-mediated phosphorylation of Bcl10. [Copyright &y& Elsevier]

Multiple kinase pathways determine serotonin transporter (SERT) regulation. We hypothesized a decrease in kinase expression with chronic selective serotonin reuptake inhibitor (SSRI) administration necessary to regulate extracellular serotonin. We studied whole brain kinase mRNA expression on Affymetrix gene chips in rats treated with placebo 3 and 21 days, fluoxetine 3 and 21 days, and citalopram 21 days. Protein kinase C (PKC)-delta, PKC-gamma, stress-activated protein kinase, cAMP-dependent protein kinase beta isoform, Janus protein kinase, and phosphofructokinase M were all down regulated chronically with citalopram and fluoxetine, but not with acute fluoxetine. The results are consistent with homeostasis of SERT function through a decrease in PK expression. [Copyright &y& Elsevier]

Tau is a microtubule-associated protein that is abnormally hyperphosphorylated in the filamentous lesions that define a number of neurodegenerative diseases collectively referred to as tauopathies. We previously showed that stress-activated protein (SAP) kinases phosphorylate tau protein at many of the hyperphosphorylated sites in vitro. Here we have developed a system to study the effects of five SAP kinases (SAPK1c/JNK1, SAPK2a/p38α, SAPK2b/p38β, SAPK3/p38γ and SAPK4/p38δ) on tau phosphorylation in intact cells. All kinases phosphorylated tau, albeit at different efficiencies. Tau was a good substrate for SAPK3/p38γ and SAPK4/p38δ, a reasonable substrate for SAPK2b/p38β and a relatively poor substrate for SAPK2a/p38α and SAPK1c/JNK1. These findings indicate that the aberrant activation of SAP kinases, especially SAPK3/p38γ and SAPK4/p38δ, could play an important role in the abnormal hyperphosphorylation of tau that is an invariant feature of the tauopathies. [Copyright &y& Elsevier]

A stress-activated protein kinase pathway comprising mitogen-activated protein kinase kinases (MKKs), c-Jun N-terminal kinase (JNK) and the transcription factor c-Jun is implicated in neuronal apoptosis. Using an immune-complex kinase assay, we measured the activation of MKK4 and MKK7 in low potassium (LK)-induced apoptosis of rat cerebellar granule neurons (CGN). MKK7, but not MKK4, was activated within the first 4–6 h in four independent sets of 14-h CGN apoptosis time-courses. CEP-1347 (500 nM), an mixed-lineage kinase 3 inhibitor, prevented MKK7 activation and cell death following exposure of CGN cultures to LK-induced apoptosis. Western blot analysis revealed that levels of phosphorylated c-Jun were elevated between 30 min and 10 h of CGN apoptosis, temporally consistent with MKK7 activation. These data suggest that MKK7 is responsible for activating the JNK pathway during LK-induced CGN apoptosis. [Copyright &y& Elsevier]

Stress-activated protein kinase (SAPK) signaling plays essential roles in eliciting adequate cellular responses to stresses and proinflammatory cytokines. SAPK pathways are composed of three successive protein kinase reactions. The phosphorylation of SAPK signaling components on Ser/Thr or Thr/Tyr residues suggests the involvement of various protein phosphatases in the negative regulation of these systems. Accumulating evidence indicates that three families of protein phosphatases, namely the Ser/Thr phosphatases, the Tyr phosphatases and the dual specificity Ser/Thr/Tyr phosphatases regulate these pathways, each mediating a distinct function. Differences in substrate specificities and regulatory mechanisms for these phosphatases form the molecular basis for the complex regulation of SAPK signaling. Here we describe the properties of the protein phosphatases responsible for the regulation of SAPK signaling pathways. [ABSTRACT FROM AUTHOR]

Two functionally different MAP kinase phosphatases (MKPs) were investigated to clarify their roles in behavioral sensitization to methamphetamine (METH). MKP-1 mRNA levels increased substantially by about 60–300% in a range of brain regions, including several cortices, the striatum and thalamus 0.5–1 h after acute METH administration. After chronic METH administration its increase was less pronounced, but a more than 50% increase was still seen in the frontal cortex. MKP-1 protein levels also increased 3 h after acute or chronic METH administration. MKP-3 mRNA levels increased by about 30–50% in several cortices, the striatum and hippocampus 1 h after acute METH administration, but only in the hippocampus CA1 after chronic METH administration. Pre-treatment with the D[sub 1] dopamine receptor antagonist, SCH23390, attenuated the METH-induced increase of MKP-1 and MKP-3 mRNA in every brain region, while pre-treatment with the NMDA receptor antagonist, MK-801, attenuated it in some regions. These findings suggest that in METH-induced sensitization, MKP-1 and MKP-3 play important roles in the neural plastic modification in widespread brain regions in the earlier induction process, but in the later maintenance process, they do so only in restricted brain regions such as MKP-1 in the frontal cortices and MKP-3 in the hippocampus. [ABSTRACT FROM AUTHOR]

Exercise has numerous growth and metabolic effects in skeletal muscle, including changes in glycogen metabolism, glucose and amino acid uptake, protein synthesis and gene transcription. However, the mechanism(s) by which exercise regulates intracellular signal transduction to the transcriptional machinery in the nucleus, thus modulating gene expression, is largely unknown. This review will provide insight on potential intracellular signalling mechanisms by which muscle contraction/exercise leads to changes in gene expression. Mitogen-activated protein kinase (MAPK) cascades are associated with increased transcriptional activity. The MAPK family members can be separated into distinct parallel pathways including the extracellular signal-regulated kinase (ERK) 1/2, the stress-activated protein kinase cascades (SAPK1/JNK and SAPK2/p38) and the extracellular signal-regulated kinase 5 (ERK5). Acute exercise elicits signal transduction via MAPK cascades in direct response to muscle contraction. Thus, MAPK pathways appear to be potential physiological mechanisms involved in the exercise-induced regulation of gene expression in skeletal muscle. [ABSTRACT FROM AUTHOR]

Hsp72, a major inducible member of the heat shock protein family, can protect cells against many cellular stresses including heat shock. In our present study, we observed that pretreatment of NIH 3T3 cells with mild heat shock (43°C for 20 min) suppressed UV-stimulated c-Jun N-terminal kinase 1 (JNK1) activity. Constitutively overexpressed Hsp72 also inhibited JNK1 activation in NIH 3T3 cells, whereas it did not affect either SEK1 or MEKK1 activity. Both in vitro binding and kinase studies indicated that Hsp72 bound to JNK1 and that the peptide binding domain of Hsp72 was important to the binding and inhibition of JNK1. In vivo binding between endogenous Hsp72 and JNK1 in NIH 3T3 cells was confirmed by coimmunoprecipitation. Hsp72 also inhibited JNK-dependent apoptosis. Hsp72 antisense oligonucteotides blocked Hsp72 production in NIH 3T3 cells in response to mild heat shock and concomitantly abolished the suppressive effect of mild heat shock on UV-induced JNK activation and apoptosis. Collectively, our data suggest strongly that Hsp72 can modulate stress-activated signaling by directly inhibiting JNK. [ABSTRACT FROM AUTHOR]

Nitric oxide modulates stretch activation of mitogen-activated protein kinases in mesangial cells. Background. In vivo, intraglomerular hypertension results in resident cell hypertrophy, proliferation and matrix protein production, leading to glomerulosclerosis. Mesangial cells (MCs) exposed to in vitro stretch also proliferate and produce matrix. We have shown activation of Jun N-terminal kinase/stress-activated protein kinase (SAPK) and p42/44 mitogen-activated protein kinase (MAPK) in stretched MCs and have also demonstrated that l-arginine decreases resident cell proliferation and protects against glomerulosclerosis in remnant kidney glomeruli, presumably by increasing nitric oxide (NO) production. Consequently, we studied whether NO could affect SAPK and p42/44 MAPK activation in stretched MCs. Methods. MCs (passages 5 to 10) cultured on type 1 collagen-coated, flexible-bottom plates were exposed to 0 to 30 minutes of cyclic strain (60 cycles per minute) by computer-driven generation of vacuum of -27 kPa, inducing 28% elongation in the diameter of the surface. Control MCs were grown on coated, flexible-bottom plates. Protein levels (by Western blot) and activity assays for SAPK/JNK and p42/44 MAPK were performed under these conditions. As maximal activation was at 10 minutes, with decay by 30 minutes, the effect of NO on kinase activation was studied at 0, 2, 5, and 10 minutes by preincubation with 70 μmol/L s-nitroso-n-acetylpenicillamine (SNAP; an NO donor) or 1 mmol/L 8-bromo cyclic guanosine monophosphate (8-bromo-cGMP). Downstream events in response to stretch and NO were studied at the time of maximal response (10 minutes) by examining nuclear translocation of SAPK with immunofluorescence microscopy and transcription factor activator protein-1 nuclear protein binding by gel mobility shift assay. The effect of kinase inhibition by NO donors on MC proliferation was studied by Western blotting for proliferating cell nuclear antigen (PCNA). Results.... [ABSTRACT FROM AUTHOR]

The responses of mitogen-activated protein kinase (MAPK) family members, including ERK (extracellular signal-regulated kinase), JNK (c-Jun NH2-terminal kinase), and p38, in the metabolic responses to whole animal freezing (up to 24 h frozen at −2.5°C) and thawing (up to 4 h at 5°C after a 12 h freeze) were examined in four organs (liver, kidney, heart, brain) of the freeze-tolerant wood frog Rana sylvatica. Levels of the active phosphorylated form of p38 increased within 20 min as an early response to freezing in liver and kidney but rose later (after 12 h) in heart. Both JNK and p38 were activated during thawing in liver, kidney and heart with temporally-distinct patterns in each organ. The only MAPK response to freeze/thaw in frog brain was a transient elevation of p38 after 90 min thawing. ERK activity did not respond to freeze/thaw in any organ. The levels of c-Fos increased during freezing in kidney and brain whereas c-Jun was unaffected by freeze/thaw. Organ-specific responses by MAPKs, particularly p38, suggest that these may have roles in regulating metabolic or gene expression responses that may be adaptive in dealing with freezing stress or metabolic recovery during thawing. [ABSTRACT FROM AUTHOR]

Background: The pathophysiology of pancreatitis and the pancreatic stress response are not well understood. In the pancreas, mitogen-activated protein kinase (MAPK) and stress-activated protein kinase (SAPK) are reportedly regulated by secretagogue stimulation and hyperstimulation. However, no data exist on the expression and regulation of pancreatic p38 Map kinase. Aims: Pancreatic expression of p38 Map kinase and MAPK, SAPK and p38 regulation during pancreatic stress were investigated. Methods: For hyperstimulation and secretory stress, cerulein was given intravenously, while hyperthermia preconditioning stress was induced by whole body hyperthermia (42°C). Results: In addition to MAPK and SAPK, p38 Map kinase was found to be expressed in the rat pancreas. Cerulein regulated all kinases time- and dose-dependently. MAPK and p38 Map kinase showed basal activity and were further activated at low cerulein doses. SAPK had no basal activity and its activation required maximal secretory to supramaximal amounts of cerulein. Cerulein hyperstimulation, inducing pancreatitis, activated p38 more rapidly than SAPK and more strongly than MAPK. In contrast to cerulein hyperstimulation stress, hyperthermia stress only activated p38 Map kinase. Conclusions: p38 Map kinase is expressed in the pancreas and is most rapidly activated following cerulein hyperstimulation. Both SAPK and p38 Map kinase are possibly important regulators during the onset of cerulein pancreatitis. [ABSTRACT FROM AUTHOR]

The role of two vertebrate mitogen-activated protein kinases (MAPKs) in mediating responses to in vivo anoxia or freezing exposures was examined in four organs (liver, heart, kidney and brain) of hatchling red-eared turtles, Trachemys scripta elegans, which are naturally tolerant of these stresses. The extracellular signal-regulated kinases were not stress-activated except in brain of frozen turtles. The c-Jun NH2-terminal kinases (JNKs) were transiently activated by anoxia exposure in all four organs (after 1 h in brain or 5 h in other organs) but activity was suppressed during freezing except in brain which showed a transient activation of JNK after 1 h. Changes in the concentrations of the transcription factors, c-Fos and c-Myc, were also stress- and organ-specific. The patterns of MAPK activation in a stress-tolerant animal suggest the relative importance of these kinase pathways in cellular adaptation to oxygen deprivation or freezing and identify novel natural activators of MAPKs in vivo. The specificity of the signaling pathways is also emphasized here as the general whole-body stresses, anoxia and freezing, activated individual MAPKs in a tissue-, time-, and stress-dependent manner. [ABSTRACT FROM AUTHOR]

The identities of the upstream activators of the mitogen-activated protein (MAP) kinase homologues termed stress-activated-protein (SAP) kinase-1 (also known as JNK or SAPK) and SAP kinase-2 (also known as p38, RK and CSBP) were investigated in rat PC12 cells and human KB cells after exposure to cellular stresses and cytokines. In PC12 cells, the same two upstream activators. SAP kinase kinase-1 (SAPKK-1) and SAPKK-2 were activated after exposure to osmotic shock, ultraviolet irradiation or the protein synthesis inhibitor anisomycin, and more weakly in response to sodium arsenite. SAPKK-1 was capable of activating both SAP kinase-1 and SAP kinase-2 and was similar, if not identical, to the previously described MAP kinase homologue MKK4. as judged by immunological criteria and by its ability to be activated by MEK kinase in vitro. In contrast, SAPKK-2 activated SAP kinase-2, but not SAP kinase-1 in vitro. In KB cells, five distinct upstream activators of SAP kinase-1 and SAP kinase-2 were induced, namely SAPKK-1, SAPKK-2, SAPKK-3, SAPKK-4 and SAPKK-5, whose appearance depended on the nature of the stimulus. SAPKK-3, which was strongly induced by every stimulus tested (osmotic shock, ultraviolet irradiation, anisomycin or IL-1), accounted for about 95% of the SAP kinase-2 activator activity in these cells, did not activate SAP kinase-1 and eluted from Mono S at a lower sail concentration than SAPKK-2, SAPKK-4 and SAPKK-5 were also eluted from Mono S at higher NaCl concentrations than SAPKK-3 and these enzymes activated SAP kinase-1 but not SAP kinase-2. SAPKK-4 was the only SAP kinase-1 activator induced by interleukin-1 or ultraviolet irradiation. while two SAP kinase-1 activators. SAPKK-1 and SAPKK-5. were induced by osmotic shock or anisomycin. SAPKK-2, SAPKK-3, SAPKK-4 and SAPKK-5 were not activated by MEK kinase in vitro, were separable from the major activator(s) of p42 MAP kinase, and were not recognised by anti-MKK4 antibodies. At least two of these enzymes are likely to be novel MAP kinase homologues. Our results demonstrate unexpected complexity in the upstream regulation of stress and cytokine-stimulated kinase cascades and indicate that the selection of the appropriate SAPKK varies with both the stimulus and the cell type. [ABSTRACT FROM AUTHOR]

Mitogen-activated protein (MAP) kinases bind tightly to many of their physiologically relevant substrates. We have identified a new subfamily of murine serine/threonine kinases, whose members, MAP kinase-interacting kinase 1 (Mnk1) and Mnk2, bind tightly to the growth factor-regulated MAP kinases, Erk1 and Erk2. Mnk1, but not Mnk2, also binds strongly to the stress-activated kinase, p38. Mnk1 complexes more strongly with inactive than active Erk, implying that Mnk and Erk may dissociate after mitogen stimulation. Erk and p38 phosphorylate Mnk1 and Mnk2, which stimulates their in vitro kinase activity toward a substrate, eukaryotic initiation factor-4E (eIF-4E). Initiation factor eIF-4E is a regulatory phosphoprotein whose phosphorylation is increased by insulin in an Erk-dependent manner. In vitro, Mnk1 rapidly phosphorylates eIF-4E at the physiologically relevant site, Ser209. In cells, Mnk1 is post-translationally modified and enzymatically activated in response to treatment with either peptide growth factors, phorbol esters, anisomycin or UV. Mitogen- and stress-mediated Mnk1 activation is blocked by inhibitors of MAP kinase kinase 1 (Mkk1) and p38, demonstrating that Mnk1 is downstream of multiple MAP kinases. Mnk1 may define a convergence point between the growth factor-activated and one of the stress-activated protein kinase cascades and is a candidate to phosphorylate eIF-4E in cells. [ABSTRACT FROM AUTHOR]

Nck, an adaptor protein composed of one SH2 and three SH3 domains, is a common target for a variety of cell surface receptors. We have identified a novel mammalian serine/threonine kinase that interacts with the SH3 domains of Nck, termed Nck Interacting Kinase (NIK). This kinase is most homologous to the Sterile 20 (Ste20) family of protein kinases. Of the members of this family, GCK and MSST1 are most similar to NIK in that they bind neither Cdc42 nor Rac and contain an N-terminal kinase domain with a putative C-terminal regulatory domain. Transient overexpression of NIK specifically activates the stressactivated protein kinase (SAPK) pathway. Both the kinase domain and C-terminal regulatory region of NIK are required for full activation of SAPK. NIK likely functions upstream of MEKK1 to activate this pathway; a dominant-negative MEK kinase 1 (MEKK1) blocks activation of SAPK by NIK. MEKK1 and NIK also associate in cells and this interaction is mediated by regulatory domains on both proteins. Two other members of this kinase family, GCK and HPK1, contain C-terminal regulatory domains with homology to that of NIK. These findings indicate that the C-terminal domain of these proteins encodes a new protein domain family and suggests that this domain couples these kinases to the SAPK pathway, possibly by interacting with MEKK1 or related kinases. [ABSTRACT FROM AUTHOR]

Extracellular stimuli such as neurotransmitters, neurotrophins, and growth factors in the brain regulate critical cellular events, including synaptic transmission, neuronal plasticity, morphological differentiation and survival. Although many such stimuli trigger Ser/Thr-kinase and tyrosine-kinase cascades, the extracellular signal-regulated kinases, ERK1 and ERK2, prototypic members of the mitogen-activated protein (MAP) kinase family, are most attractive candidates among protein kinases that mediate morphological differentiation and promote survival in neurons. ERK1 and ERK2 are abundant in the central nervous system (CNS) and are activated during various physiological and pathological events such as brain ischemia and epilepsy. In cultured hippocampal neurons, simulation of glutamate receptors can activate ERK signaling, for which elevation of intracellular Ca2+ is required. In addition, brain-derived neurotrophic factor and growth factors also induce the ERK signaling and here, receptor-coupled tyrosine kinase activation has an association. We describe herein intracellular cascades of ERK signaling through neurotransmitters and neurotrophic factors. Putative functional implications of ERK and other MAP-kinase family members in the central nervous system are give attention. [ABSTRACT FROM AUTHOR]

Two distinct stress-activated protein kinase (JNKa and b) cDNAs were isolated from a carp ovary cDNA library. These cDNAs contained a full-length open reading frame encoding 427 amino acid residues with a predicted mass of 48.6 kDa. The deduced amino acid sequences of JNKa and b were 95.8% identical, with 18 residues replaced, and showed a high degree of sequence similarity to mammalian JNK/SAPK subgroup including the common dual phosphorylation motif of TPY. By Northern blot analysis, the carp JNKs were found to be abundant in the brain and ovary. Detailed study by RT-PCR assay revealed ubiquitous expression of JNKb, although expression of JNKa was specific to the brain and ovary. The high level expression of both JNKa and b in the ovary implies that JNKs play an important role in egg maturation or ectogenetic early development. [ABSTRACT FROM AUTHOR]