Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.7.11.24 (mitogen-activated protein kinase)
 95,810 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The activation of human mitogen-activated protein kinase kinase 1 (MKK1) is achieved by phosphorylation at Ser218 and Ser222 within a regulatory loop. Partial activation was achieved by replacing these residues with aspartic/glutamic acid. Higher activity was obtained by introducing four acidic residue substitutions in the regulatory loop, indicating that acidic residues in the loop stabilize an active configuration by the introduction of negative charge. Activation of MKK1 is also achieved by deleting residues 44-51, N-terminal to the consensus catalytic core. Although substitution of residues within this segment by alanine does not affect activity, introduction of proline residues elevates kinase activity, indicating that activation results from perturbation of secondary structure within residues 44-51. Pseudosubstrate inhibition, a commonly observed mechanism of kinase regulation, is not operative in this process. Both the acidic substitutions and the N-terminal deletion increase Vmax, V/K(m),ERK2, and V/K(m),ATP, as is also observed following phosphorylation of wild-type MKK1. A synergistic enhancement of these steady-state rate parameters occurs upon combining the mutations, suggesting that conformational changes induced by mutagenesis together mimic those seen upon phosphorylation.
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PMID:Interdependent domains controlling the enzymatic activity of mitogen-activated protein kinase kinase 1. 895 7

The Fas receptor mediates a signalling cascade resulting in programmed cell death (apoptosis) within hours of receptor cross-linking. In this study Fas activated the stress-responsive mitogen-activated protein kinases, p38 and JNK, within 2 h in Jurkat T lymphocytes but not the mitogen-responsive kinase ERK1 or pp70S6k. Fas activation of p38 correlated temporally with the onset of apoptosis, and transfection of constitutively active MKK3 (glu), an upstream regulator of p38, potentiated Fas-induced cell death, suggesting a potential involvement of the MKK3/p38 activation pathway in Fas-mediated apoptosis. Fas has been shown to require ICE (interleukin-1 beta-converting enzyme) family proteases to induce apoptosis from studies utilizing the cowpox ICE inhibitor protein CrmA, the synthetic tetrapeptide ICE inhibitor YVAD-CMK, and the tripeptide pan-ICE inhibitor Z-VAD-FMK. In this study, crmA antagonized, and YVAD-CMK and Z-VAD-FMK completely inhibited, Fas activation of p38 kinase activity, demonstrating that Fas-dependent activation of p38 requires ICE/CED-3 family members and conversely that the MKK3/p38 activation cascade represents a downstream target for the ICE/CED-3 family proteases. Intriguingly, p38 activation by sorbitol and etoposide was resistant to YVAD-CMK and Z-VAD-FMK, suggesting the existence of an additional mechanism(s) of p38 regulation. The ICE/CED-3 family-p38 regulatory relationship described in the current work indicates that in addition to the previously described destructive cleavage of substrates such as poly(ADP ribose) polymerase, lamins, and topoisomerase, the apoptotic cysteine proteases also function to regulate stress kinase signalling cascades.
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PMID:Fas activation of the p38 mitogen-activated protein kinase signalling pathway requires ICE/CED-3 family proteases. 897 82

The 78-kDa protein kinase Mekk1 plays an important role in the stress response pathway that involves the activation of downstream kinases Sek1 and stress-activated protein kinase/c-Jun NH2-terminal kinase. Conserved serine and threonine residues located between the kinase subdomains VII and VIII of many protein kinases are phosphorylated for maximal kinase activation. Two threonine residues within this region in Mekk1 at positions 560 and 572, but not the serine at 557, were shown to be essential for catalytic activity in this study. When these threonine residues were replaced with alanine, there was a significant loss in phosphotransferase activity toward the primary substrate, Sek1, and a large decrease in autophosphorylation activity. Site-directed mutagenesis demonstrated that these threonine residues cannot be replaced with either serine or glutamic acid for preservation of phosphotransferase activity. Further examination of the Mekk1 mutants isolated from 32P-labeled transfected COS cells showed that Thr-560 and Thr-572 were indeed phosphorylated after two-dimensional tryptic-chymotryptic phosphopeptide analysis. Additional determinants in the NH2-terminal domain of Mekk1 also play a role in the regulation of Mekk1 activity. Although Pak3 and PKC can activate Mekk1 in vivo, this interaction is indirect and independent, since there was no direct phosphorylation of Mekk1 by Pak3 or PKC or of Pak3 by PKC, respectively.
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PMID:Identification of two essential phosphorylated threonine residues in the catalytic domain of Mekk1. Indirect activation by Pak3 and protein kinase C. 906 12

The small GTP-binding Rho proteins control a variety of biological activities, including organization of the actin cytoskeleton, regulation of gene expression and cellular transformation. In contrast, Ras proteins do not induce actin stress fibers, but potently transform cells which exhibit a morphology clearly distinct from that caused by activated forms of Rho. To investigate whether nuclear signaling and oncogenic potential of Rho are a consequence of its profound effect on cytoskeletal organization, we replaced each amino acid in the Rho effector loop with those of Ras, or replaced conserved residues with others known to result in differential signaling capability when introduced into Ras and Rac1. These Rho mutants did not gain the ability to induce the MAPK, JNK or p38 pathways but, surprisingly, all Rho effector loop mutants still continued to induce actin stress fiber formation. However, three of these Rho mutants, with substitutions of leucine-39, glutamic acid-39, or cysteine-42, lost the ability to stimulate gene transcription via the serum response factor (SRF) and failed to induce neoplastic transformation. Thus, these results indicate that cytoskeletal changes are not sufficient to induce the transformed phenotype, and that Rho-effector molecules regulating the actin cytostructure are distinct from those signaling to the nucleus and subverting normal growth control.
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PMID:Effector domain mutants of Rho dissociate cytoskeletal changes from nuclear signaling and cellular transformation. 974 78

To address a role of mitogen-activated protein kinase (MAPK) in the regulation of glucose transport, we made a constitutively active mutant of MAPK kinase (MAPKK) and introduced it into 3T3-L1 preadipocytes by using a retrovirus-mediated transfection procedure. The deletion of 20 amino acids (those between and including 32 and 51) in the amino terminal region of Xenopus MAPKK and the replacement of serine residues on the 218 and 222 positions by glutamic acid (dSESE-MAPKK) let Xenopus MAPKK constitutively active. The isolated cell clones differently expressing dSESE-MAPKK (clone 219 higher expression, clone 233 lower expression) efficiently differentiated to adipocytes by a standard differentiation cocktail. Accordingly, the increased expression of dSESE-MAPKK protein during differentiation resulted in the increased basal MAPK activity in clone 219 adipocytes and, to a lesser extent, in clone 233 adipocytes. In contrast to clone 233 and parental adipocytes, basal 2-deoxyglucose uptake was enhanced fourfold in clone 219 adipocytes, in accordance with increased expression of GLUT1 mRNA and protein. Whereas GLUT4 mRNA was similarly expressed in all of the adipocytes, GLUT4 protein appeared to decrease in clone 219 adipocytes. More importantly, subcellular fractionation studies showed that the localization of both GLUT1 and GLUT4 in the plasma membranes (PMs) was markedly increased in the basal state in clone 219 adipocytes compared with that in clone 233 and parental adipocytes, in which both glucose transporters were preferentially located in intracellular compartments. Consequently, insulin-induced translocation of GLUT1 was abolished in clone 219 adipocytes, although the remaining intracellular GLUT4 was still responsive to insulin stimulation, which led to the movement to the PM. As combined effects on the situation of GLUT1 and GLUT4, the foldness of insulin stimulation of glucose transport based on the basal activity was reduced in cells expressing constitutively active MAPKK. These results imply that chronic activation of MAPK could be one of the mechanisms for insulin resistance.
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PMID:Constitutively active mitogen-activated protein kinase kinase increases GLUT1 expression and recruits both GLUT1 and GLUT4 at the cell surface in 3T3-L1 adipocytes. 1086 53

To characterize the differentiation events that selectively target insulin-producing cells to interleukin (IL)-1beta-induced apoptosis, we studied IL-1beta signaling via mitogen-activated protein kinase (MAPK) and stress-activated protein kinase in 2 pancreatic endocrine cell lines. We studied the glucagon-secreting AN-glu cell line and the insulin and the islet amyloid polypeptide-producing beta-cell line (AN-ins cells), which is derived by stable transfection of AN-glu cells with the transcription factor pancreatic duodenal homeobox factor-1. AN-ins cells were more sensitive to the cytotoxic action of IL-1beta. This increased sensitivity was not associated with a more pronounced IL-l-induced nitric oxide production in AN-ins cells, but it correlated with a more marked activation of the 3 MAPKs extracellular signal-regulated kinases (ERKs)-1/2, c-Jun NH2-terminal kinase (JNK), and p38 MAPK (p38). This led to increased phosphorylation of the transcription factors c-Jun, Elk-1, and ATF2 and of heat shock protein 25. Inhibition of ERK-1/2 and p38 did not prevent but aggravated IL-1beta-induced cell death. In contrast, inhibition of JNK by transfection with the dominant negative inhibitor of the JNK-binding domain prevented apoptosis in both cell types. Cell death could be elicited by overexpressing the catalytic domain of MAPK kinase kinase 1, a specific activator of JNK and nuclear factor-kappaB, which does not recruit ERK-1/2 or p38. Coactivation of ERK-1/2 with JNK did not prevent apoptosis. In conclusion, increased MAPK signaling in response to IL-1beta may represent a novel molecular marker of beta-cell differentiation. JNK inhibition represents an effective means of preventing IL-1beta-activated beta-cell destruction.
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PMID:The c-Jun amino-terminal kinase pathway is preferentially activated by interleukin-1 and controls apoptosis in differentiating pancreatic beta-cells. 1096 30

The small G-protein Ras, a critical component in the signalling pathways regulating cell growth, is involved in the tonic upregulation of voltage-dependent calcium channels (VDCCs) in rat sensory neurones. To investigate which downstream effector(s) of Ras is involved in this process, a series of Ras mutant cDNAs were co-expressed with green fluorescent protein (GFP) in primary cultured rat dorsal root ganglion neurones (DRGs). Constitutively active V12Ras (glycine 12 to valine) markedly increased basal calcium current density by 41 % compared with control cells (GFP alone). In contrast, a farnesylation-defective mutant, V12S186Ras (cysteine 186 to serine; activates no downstream effectors), significantly reduced calcium current density by 47 %. Ras effector region mutants V12C40 (tyrosine 40 to cysteine; activates the p110 alpha-subunit of phosphatidylinositol 3-kinase) and V12G37 (glutamic acid 37 to glycine; activates Ral guanine nucleotide dissociation stimulator) had no significant effect on VDCC current. However, V12S35Ras (threonine 35 to serine; activates Raf-1 and the mitogen-activated protein kinase (MAPK) pathway) markedly increased basal calcium current density by 67 %, suggesting that Raf-1 activation is sufficient for Ras enhancement of calcium current in these cells. Raf-1 activates MEK (MAPK kinase) in the MAPK pathway, and the MEK inhibitor U0126 reduced calcium current by 45 % after 10-15 min, whereas the inactive analogue U0124 had no effect. This rapid time course for MEK inhibition suggests direct modulation of VDCCs via the Ras-MAPK pathway rather than gene expression-mediated effects. The relative proportions of omega-conotoxin GVIA- and nicardipine-sensitive N- ( approximately 40 %) and L- ( approximately 40 %) type currents were unaffected by either V12S35Ras expression or U0126 pre-treatment, suggesting that all components of calcium current in DRGs, are enhanced via this pathway.
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PMID:Regulation of voltage-dependent calcium channels in rat sensory neurones involves a Ras-mitogen-activated protein kinase pathway. 1099 May 31

The activation of both phosphatidylinositol 3-kinase (PI3-kinase) and p38 mitogen-activated protein kinase (p38 MAPK) is required for muscle differentiation. However, it is not known whether the signals from these two kinases interact during this process. In this work, we have investigated this using H9c2 cardiac myoblasts. The p38 MAPK-specific inhibitor SB203580 blocked muscle differentiation and suppressed the expression of myogenin and myosin heavy chain in a concentration-dependent manner. Consistent with this, expression of a wild-type p38 MAPK (Ha-p38) or a constitutively active MAPK kinase 6 (MKK6(glu)) promoted the rate of differentiation into multinucleated myotubes. LY294002, a PI3-kinase inhibitor, suppressed in a dose-dependent manner not only muscle differentiation but also activation of p38 MAPK. In addition, expression of a constitutively active form of PI3-kinase (p110*) enhanced myotube formation and p38 MAPK activation, while expression of a dominant negative form of PI3-kinase (Deltap85) attenuated these responses. Furthermore, SB203580 suppressed differentiation of H9c2 cells expressing p110*. Interestingly, LY294002 also suppressed differentiation of H9c2 cells expressing Ha-p38 or MKK6(glu). However, SB203580 did not affect PI3-kinase activity, suggesting that PI3-kinase myogenic signaling to p38 MAPK is unidirectional. Taken together, we concluded that PI3-kinase activates p38 MAPK, which in turn stimulates muscle differentiation, but that p38 MAPK does not substitute for PI3-kinase in this process.
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PMID:Phosphatidylinositol 3-kinase stimulates muscle differentiation by activating p38 mitogen-activated protein kinase. 1102 4

Ras p21 signaling is involved in multiple aspects of growth, differentiation, and stress response [1-2]. There is evidence pointing to superoxides as relays of Ras signaling messages. Chemicals with antioxidant activity suppress Ras-induced DNA synthesis. The inhibition of Ras significantly reduces the production of superoxides by the NADPH-oxidase complex [3]. Kirsten and Harvey are nonallelic Ras cellular genes that share a high degree of structural and functional homology. The sequences of Ki- and Ha-Ras proteins are almost identical. They diverge only in the 20-amino acid hypervariable domain at the COOH termini. To date, their functions remain indistinguishable [4]. We show that Ki- and Ha-Ras genes differently regulate the redox state of the cell. Ha-Ras-expressing cells produce high levels of reactive oxygen species (ROS) by inducing the NADPH-oxidase system. Ki-Ras, on the other hand, stimulates the scavenging of ROS by activating posttranscriptionally the mitochondrial antioxidant enzyme, Mn-superoxide dismutase (Mn-SOD), via an ERK1/2-dependent pathway. Glutamic acid substitution of the four lysine residues in the polybasic stretch at the COOH terminus of Ki-Ras completely abolishes the activation of Mn-SOD, although it does not inhibit ERK1/2-induced transcription. In contrast, an alanine substitution of the cysteine of the CAAX box has very little effect on Mn-SOD activity but eliminates ERK1/2- dependent transcription.
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PMID:Opposing functions of Ki- and Ha-Ras genes in the regulation of redox signals. 1136 7

Anthrax is primarily a disease of herbivores caused by gram-positive, aerobic, spore-forming Bacillus anthracis. Humans are accidental hosts through the food of animal origin and animal products. Anthrax is prevelant in most parts of the globe, and cases of anthrax have been reported from almost every country. Three forms of the disease have been recognized: cutaneous (through skin), gastrointestinal (through alimentary tract), and pulmonary (by inhalation of spores). The major virulence factors of Bacillus anthracis are a poly-D glutamic acid capsule and a three-component protein exotoxin. The genes coding for the toxin and the enzymes responsible for capsule production are carried on plasmid pXO1 and pXO2, respectively. The three proteins of the exotoxin are protective antigen (PA, 83 kDa), lethal factor (LF, 90 kDa), and edema factor (EF, 89 kDa). The toxins follow the A-B model with PA being the B moeity and LF/EF, the alternative A moeities. LF and EF are individually nontoxic, but in combination with PA form two toxins causing different pathogenic responses in animals and cultured cells. PA + LF forms the lethal toxin and PA + EF forms the edema toxin. During the process of intoxication, PA binds to the cell surface receptor and is cleaved at the sequence RKKR (167) by cell surface proteases such as furin generating a cell-bound, C-terminal 63 kDa protein (PA63). PA63 possesses a binding site to which LF or EF bind with high affinity. The complex is then internalized by receptor-mediated endocytosis. Acidification of the vesicle leads to instertion of PA63 into the endosomal membrane and translocation of LF/EF across the bilayer into the cytosol where they exert their toxic effects. EF has a calcium- and calmodulin-dependent adenylate cyclase activity. Recent reports indicate that LF is a protease that cleaves the amino terminus of mitogen-activated protein kinase kinases 1 and 2 (MAPKK1 and 2), and this cleavage inactivates MAPKK1 and thus inhibits the mitogen-activated protein kinase signal transduction pathway. We describe in detail the studies so far done on unraveling the molecular mechanisms of pathogenesis of Bacillus anthracis.
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PMID:Anthrax toxin. 1159 78


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