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M Kracht

Publications and source records attributed to M Kracht.

At least 19 recordsLinked to original sources
PubMed2006

The mitogen-activated protein kinase (MAPK)-activated protein kinases MK2 and MK3 cooperate in stimulation of tumor necrosis factor biosynthesis and stabilization of p38 MAPK.

MK2 and MK3 represent protein kinases downstream of p38 mitogen-activated protein kinase (MAPK). Deletion of the MK2 gene in mice resulted in an impaired inflammatory response although MK3, which displays extensive structural similarities and identical functional properties in vitro, is still present. Here, we analyze tumor necrosis factor (TNF) production and expression of p38 MAPK and tristetraprolin (TTP) in MK3-deficient mice and demonstrate that there are no significant differences with wild-type animals. We show that in vivo MK2 and MK3 are expressed and activated in parallel. However, the level of activity of MK2 is always significantly higher than that of MK3. Accordingly, we hypothesized that MK3 could have significant effects only in an MK2-free background and generated MK2/MK3 double-knockout mice. Unexpectedly, these mice are viable and show no obvious defects due to loss of compensation between MK2 and MK3. However, there is a further reduction of TNF production and expression of p38 and TTP in double-knockout mice compared to MK2-deficient mice. This finding, together with the observation that ectopically expressed MK3 can rescue MK2 deficiency similarly to MK2, indicates that both kinases share the same physiological function in vivo but are expressed to different levels.

Animals↗
PubMed2005

Alpha-toxin is produced by skin colonizing Staphylococcus aureus and induces a T helper type 1 response in atopic dermatitis.

BACKGROUND: Staphylococcus aureus is a well known trigger factor of atopic dermatitis (AD). Besides the superantigens, further exotoxins are produced by S. aureus and may have an influence on the eczema. OBJECTIVE: To explore the impact of staphylococcal alpha-toxin on human T cells, as those represent the majority of skin infiltrating cells in AD. METHODS: Adult patients with AD were screened for cutaneous colonization with alpha-toxin producing S. aureus. As alpha-toxin may induce necrosis, CD4(+) T cells were incubated with sublytic alpha-toxin concentrations. Proliferation and up-regulation of IFN-gamma on the mRNA and the protein level were assessed. The induction of t-bet translocation in CD4(+) T cells was detected with the Electrophoretic Mobility Shift Assay. RESULTS: Thirty-four percent of the patients were colonized with alpha-toxin producing S. aureus and alpha-toxin was detected in lesional skin of these patients by immunohistochemistry. Sublytic alpha-toxin concentrations induced a marked proliferation of isolated CD4(+) T cells. Microarray analysis indicated that alpha-toxin induced particularly high amounts of IFN-gamma transcripts. Up-regulation of IFN-gamma was confirmed both on the mRNA and the protein level. Stimulation of CD4(+) T cells with alpha-toxin resulted in DNA binding of t-bet, known as a key transcription factor involved into primary T helper type 1 (Th1) commitment. CONCLUSION: alpha-toxin is produced by S. aureus isolated from patients with AD. We show here for the first time that sublytic alpha-toxin concentrations activate T cells in the absence of antigen-presenting cells. Our results indicate that alpha-toxin is relevant for the induction of a Th1 like cytokine response. In AD, this facilitates the development of Th1 cell dominated chronic eczema.

Adult↗
PubMed2001

Influenza virus-induced AP-1-dependent gene expression requires activation of the JNK signaling pathway.

Influenza A virus infection of cells results in the induction of a variety of antiviral cytokines, including those that are regulated by transcription factors of the activating protein-1 (AP-1) family. Here we show that influenza virus infection induces AP-1-dependent gene expression in productively infected cells but not in cells that do not support viral replication. Among the AP-1 factors identified to bind to their cognate DNA element during viral infections of Madin-Darby canine kidney and U937 cells are those that are regulated via phosphorylation by JNKs. Accordingly, we observed that induction of AP-1-dependent gene expression correlates with a strong activation of JNK in permissive cells, which appears to be caused by viral RNA accumulation during replication. Blockade of JNK signaling at several levels of the cascade by transient expression of dominant negative kinase mutants and inhibitory proteins resulted in inhibition of virus-induced JNK activation, reduced AP-1 activity, and impaired transactivation of the IFN-beta promoter. Virus yields from transfected and infected cells in which JNK signaling was inhibited were higher compared with the levels from control cells. Therefore, we conclude that virus-induced activation of JNK and AP-1 is part of the innate antiviral response of the cell.

Activating Transcription Factor 2↗
PubMed2001

Influenza virus-induced AP-1-dependent gene expression requires activation of the JNK signaling pathway.

Influenza A virus infection of cells results in the induction of a variety of antiviral cytokines, including those that are regulated by transcription factors of the activating protein-1 (AP-1) family. Here we show that influenza virus infection induces AP-1-dependent gene expression in productively infected cells but not in cells that do not support viral replication. Among the AP-1 factors identified to bind to their cognate DNA element during viral infections of Madin-Darby canine kidney and U937 cells are those that are regulated via phosphorylation by JNKs. Accordingly, we observed that induction of AP-1-dependent gene expression correlates with a strong activation of JNK in permissive cells, which appears to be caused by viral RNA accumulation during replication. Blockade of JNK signaling at several levels of the cascade by transient expression of dominant negative kinase mutants and inhibitory proteins resulted in inhibition of virus-induced JNK activation, reduced AP-1 activity, and impaired transactivation of the IFN-beta promoter. Virus yields from transfected and infected cells in which JNK signaling was inhibited were higher compared with the levels from control cells. Therefore, we conclude that virus-induced activation of JNK and AP-1 is part of the innate antiviral response of the cell.

Animals↗
PubMed2001

I kappa B-independent control of NF-kappa B activity by modulatory phosphorylations.

Activation of the transcription factor nuclear factor kappa B (NF-kappa B) requires its release from inhibitor of NF-kappa B (I kappa B) proteins in the cytoplasm. Much work has focussed on the identification of pathways regulating this cytosolic rate-limiting step of NF-kappa B activation. However, there is increasing evidence for another complex level of NF-kappa B activation, which involves modulatory phosphorylations of the DNA-binding subunits. These phosphorylations can control several functions of NF-kappa B, including DNA binding and transactivation properties, as well as interactions between the transcription factor and regulatory proteins. Although their overall impact on NF-kappa B function has yet to be determined, modifications of this factor will very probably provide a mechanism to fine tune NF-kappa B function.

Binding Sites↗
PubMed2000

The NF-kappa b repressing factor is involved in basal repression and interleukin (IL)-1-induced activation of IL-8 transcription by binding to a conserved NF-kappa b-flanking sequence element.

Interleukin (IL)-8, a prototypic chemokine, is rapidly induced by the pro-inflammatory cytokine IL-1 but is barely detectable in noninduced cells. Although there is clear evidence that the transcription factor NF-kappaB plays a central role in inducible IL-8 transcription, very little is known about the cis-elements and trans-acting factors involved in silencing of the IL-8 promoter. By sequence comparison with the interferon-beta promoter, we found a negative regulatory element (NRE) in the IL-8 promoter overlapping partially with the NF-kappaB response element. Here we show that an NF-kappaB-repressing factor (NRF) binds to the IL-8 promoter NF-kappaB-NRE. Reduction of cellular NRF by expressing NRF antisense RNA results in spontaneous IL-8 gene expression. In contrast, IL-1-induced IL-8 secretion is strongly impaired by expressing NRF antisense RNA. Mutation of the NRE site results in loss of NRF binding and increased basal IL-8 transcription. On the other hand IL-1-induced IL-8 transcription is decreased by mutating the NRE. These data provide evidence for a dual role of the NRF in IL-8 transcription. Although in the absence of stimulation it is involved in transcriptional silencing, in IL-1-induced cells it is required for full induction of the IL-8 promoter.

Base Sequence↗
PubMed2000

The MAPK kinase kinase TAK1 plays a central role in coupling the interleukin-1 receptor to both transcriptional and RNA-targeted mechanisms of gene regulation.

Mechanisms of fulminant gene induction during an inflammatory response were investigated using expression of the chemoattractant cytokine interleukin-8 (IL-8) as a model. Recently we found that coordinate activation of NF-kappaB and c-Jun N-terminal protein kinase (JNK) is required for strong IL-8 transcription, whereas the p38 MAP kinase (MAPK) pathway stabilizes the IL-8 mRNA. It is unclear how these pathways are coupled to the receptor for IL-1, an important physiological inducer of IL-8. Expression of the MAP kinase kinase kinase (MAPKKK) TAK1 together with its coactivator TAB1 in HeLa cells activated all three pathways and was sufficient to induce IL-8 formation, NF-kappaB + JNK2-mediated transcription from a minimal IL-8 promoter, and p38 MAPK-mediated stabilization of a reporter mRNA containing IL-8-derived regulatory mRNA sequences. Expression of a kinase-inactive mutant of TAK1 largely blocked IL-1-induced transcription and mRNA stabilization, as well as formation of endogenous IL-8. Truncated TAB1, lacking the TAK1 binding domain, or a TAK1-derived peptide containing a TAK1 autoinhibitory domain were also efficient in inhibition. These data indicate that the previously described three-pathway model of IL-8 induction is operative in response to a physiological stimulus, IL-1, and that the MAPKKK TAK1 couples the IL-1 receptor to both transcriptional and RNA-targeted mechanisms mediated by the three pathways.

Adaptor Proteins, Signal Transducing↗
PubMed1999

The p38 MAP kinase pathway signals for cytokine-induced mRNA stabilization via MAP kinase-activated protein kinase 2 and an AU-rich region-targeted mechanism.

Stabilization of mRNAs contributes to the strong and rapid induction of genes in the inflammatory response. The signaling mechanisms involved were investigated using a tetracycline-controlled expression system to determine the half-lives of interleukin (IL)-6 and IL-8 mRNAs. Transcript stability was low in untreated HeLa cells, but increased in cells expressing a constitutively active form of the MAP kinase kinase kinase MEKK1. Destabilization and signal-induced stabilization was transferred to the stable beta-globin mRNA by a 161-nucleotide fragment of IL-8 mRNA which contains an AU-rich region, as well as by defined AU-rich elements (AREs) of the c-fos and GM-CSF mRNAs. Of the different MEKK1-activated signaling pathways, no significant effects on mRNA degradation were observed for the SAPK/JNK, extracellular regulated kinase and NF-kappaB pathways. Selective activation of the p38 MAP kinase (=SAPK2) pathway by MAP kinase kinase 6 induced mRNA stabilization. A dominant-negative mutant of p38 MAP kinase interfered with MEKK1 and also IL-1-induced stabilization. Furthermore, an active form of the p38 MAP kinase-activated protein kinase (MAPKAP K2 or MK2) induced mRNA stabilization, whereas a negative interfering MK2 mutant interfered with MAP kinase kinase 6-induced stabilization. These findings indicate that the p38 MAP kinase pathway contributes to cytokine/stress-induced gene expression by stabilizing mRNAs through an MK2-dependent, ARE-targeted mechanism.

3' Untranslated Regions↗
PubMed1999

Molecular mechanisms of T lymphocyte activation: convergence of T cell antigen receptor and IL-1 receptor-induced signaling at the level of IL-2 gene transcription.

Co-stimulation of murine EL-4 thymoma cells-carrying high numbers of TCR and type I IL-1 receptors (IL-1R)-with anti-CD3 antibodies and IL-1 resulted in synergistic enhancement of IL-2 synthesis. While the extracellular signal-regulated kinase (ERK) cascade was activated by both receptors, IL-1 preferentially stimulated Jun-N-terminal kinases (JNK) and p38 mitogen-activated kinase or microtubule-associated protein kinase (MAPK). Interruption of TCR- or IL-1R-stimulated ERK cascade by PD-98059, a specific inhibitor of MAP/ERK kinase (MEK), resulted in partial suppression of nuclear factor of activated T cells activation and in complete inhibition of IL-1-stimulated NFkappaB activation. Suppression of activation of both MEK and p38 MAPK resulted in significant inhibition of IL-2 gene expression. The results show that maximal activation of the IL-2 gene requires activation of at least two different protein kinase cascades, i.e. of the ERK and p38 pathways but presumably also that of JNK which converge at the level of the IL-2 promoter resulting in enhancement of its transcriptional activity.

Animals↗
PubMed1999

Induction of interleukin-8 synthesis integrates effects on transcription and mRNA degradation from at least three different cytokine- or stress-activated signal transduction pathways.

A hallmark of inflammation is the burst-like formation of certain proteins, initiated by cellular stress and proinflammatory cytokines like interleukin 1 (IL-1) and tumor necrosis factor, stimuli which simultaneously activate different mitogen-activated protein (MAP) kinases and NF-kappaB. Cooperation of these signaling pathways to induce formation of IL-8, a prototype chemokine which causes leukocyte migration and activation, was investigated by expressing active and inactive forms of protein kinases. Constitutively active MAP kinase kinase 7 (MKK7), an activator of the stress-activated protein kinase/c-Jun N-terminal kinase (SAPK/JNK) pathway, induced IL-8 synthesis and transcription from a minimal IL-8 promoter. Furthermore, MKK7 synergized in both effects with NF-kappaB-inducing kinase (NIK). Activation of the IL-8 promoter by either of the kinases required functional NF-kappaB and AP-1 sites. While NIK and MKK7 did not affect degradation of IL-8 mRNA, an active form of MKK6, which selectively activates p38 MAP kinase, induced marked stabilization of the transcript and further increased IL-8 protein formation induced by NIK plus MKK7. Consistently, the MAP kinase kinase kinase MEKK1, which can activate NF-kappaB, SAPK/JNK, and p38 MAP kinases, most potently induced IL-8 formation. These results provide evidence that maximal IL-8 gene expression requires the coordinate action of at least three different signal transduction pathways which cooperate to induce mRNA synthesis and suppress mRNA degradation.

Binding Sites↗
PubMed1999

Antiviral and hemolytic activities of surfactin isoforms and their methyl ester derivatives.

Inactivation of enveloped viruses (VSV, SFV, and SHV-1) by surfactin lipopeptides was dependent on the hydrophobicity, i.e. the number of carbon atoms of the fatty acid, and on the charge of the peptide moiety as well as on the virus species. Surfactins with fatty acid chains of 13 carbon atoms showed very low antiviral activity in comparison to C14 and C15 isoforms. C15 surfactin monomethyl ester also inactivated SFV which was resistant to the mixture of surfactin isoforms as produced by Bacillus subtilis. In contrast, the dimethyl ester showed no virus-inactivation capacity. Disintegration of viral structures as determined by electron microscopy after inactivation of VSV and SFV was comparable to the titer reduction. The effect of the surfactin isoforms and methyl esters on erythrocyte hemolysis correlated with the virus-inactivation capacity. Surfactins with a fatty acid chain moiety of 15 carbon atoms and one negative charge showed the highest antiviral activity.

Animals↗
PubMed1998

Stress-activated protein kinase/Jun N-terminal kinase is required for interleukin (IL)-1-induced IL-6 and IL-8 gene expression in the human epidermal carcinoma cell line KB.

The cytokine interleukin-1 (IL-1) is a major inflammatory hormone which activates a broad range of genes during inflammation. The signaling mechanisms triggered by IL-1 include activation of several distinct protein kinase systems. The stress-activated protein kinase (SAPK), also termed Jun N-terminal kinase (JNK), is activated particularly strongly by the cytokine. In an attempt to delineate its role in activation of gene expression by IL-1, we inhibited the IL-1-induced SAPK/JNK activity by stable overexpression of either a catalytically inactive mutant of SAPKbeta (SAPKbeta(K-R)) or antisense RNA to SAPKbeta in human epidermal carcinoma cells. A detailed analysis of signal transduction in those cells showed that activation of neither NFkappaB nor p38 mitogen-activated protein kinase was affected, suggesting that we achieved specific blockade of the SAPK/JNK. In untransfected and vector-transfected KB cells, IL-1 induced a strong increase in expression of IL-6 and IL-8 mRNA, along with the synthesis of high amounts of the proteins. In two KB cell clones stably overexpressing the mutant SAPKbeta(K-R), and three clones stably overexpressing antisense RNA to SAPKbeta, expression of IL-6 and IL-8 in response to IL-1 was strongly reduced at both the mRNA and protein level. These data indicate that the SAPK/JNK pathway provides an indispensable signal for IL-1-induced expression of IL-6 and IL-8.

Calcium-Calmodulin-Dependent Protein Kinases↗
PubMed1997

Selective activation of JNK/SAPK by interleukin-1 in rabbit liver is mediated by MKK7.

Activation of jun N-terminal kinase (JNK)/stress-activated protein kinase (SAPK) by interleukin-1 (IL-1) has been reported in many cells and in rabbit liver. Here we report selective activation of JNK/SAPK, without activation of p38 or p42 mitogen-activated protein kinases (MAPKs), by IL-1 in rabbit liver. We identified an IL-1 regulated JNK/SAPK activator present in rabbit liver using S Sepharose chromatography. It was purified and immunoprecipitated by two antisera to MAP kinase kinase 7 (MKK7). It was not recognised by an antibody to MKK4. We conclude that MKK7 is the activator of JNK/SAPK activated by IL-1 in liver and that JNK/SAPK is the only MAPK activated by IL-1 in liver.

Amino Acid Sequence↗
PubMed1997

The interleukin-1 receptor accessory protein (IL-1RAcP) is essential for IL-1-induced activation of interleukin-1 receptor-associated kinase (IRAK) and stress-activated protein kinases (SAP kinases).

Interleukin-1 (IL-1) is a central mediator of the immune system involved in acute and chronic inflammatory responses. Although the sequences of two types of IL-1 receptors are known, the exact molecular events resulting in signal transduction and coupling to downstream signaling elements remain unclear. The recently cloned IL-1 receptor accessory protein (IL-1RAcP) has been suggested as a co-receptor molecule for IL-1RI, supported by the observation that its expression correlates to IL-1 responsiveness. We transfected the EL-4 subline D6/76 with IL-1RAcP cDNA. This cell line is an IL-1 non-responder expressing IL-1RI but lacking constitutive IL-1RAcP expression. The expression of IL-1RAcP in EL-4 D6/76 was sufficient to restore IL-1-induced activation of interleukin-1 receptor-associated kinase and of stress-activated protein kinases, translocation of the transcription factors NFkappaB and IL-1 NF to the nucleus, and induction of IL-2 mRNA synthesis. These results proved that IL-1RAcP is an indispensible molecule in the IL-1 receptor signal transduction complex, necessary to link events on the plasma membrane level to downstream signaling pathways, allowing IL-1-dependent activation of transcription factors and gene expression.

Animals↗
PubMed1996

Interleukin 1 activates jun N-terminal kinases JNK1 and JNK2 but not extracellular regulated MAP kinase (ERK) in human glomerular mesangial cells.

Interleukin 1 (IL-1) potently activates human glomerular mesangial cells (HMC). In cytosolic extracts of IL-1-stimulated HMC or in anion exchange chromatography fractions we could not find any change in phosphorylation of myelin basic protein (MBP), a good substrate for extracellular regulated kinase (ERK). In contrast, IL-1 stimulated GST-jun kinase activity at least 10-fold. The jun kinase activity could be characterised as JNK1 and JNK2 at the protein and mRNA level. IL-1, TNF, UV light and osmotic stress, but not PMA, LPS, IL-3, IL-4, IL-6, IL-8, IL-10, IL-13, GM-CSF, PDGF, bFGF, TGF-beta and interferon-gamma were able to stimulate jun kinase activity in HMC, suggesting that jun kinase is selectively mediating signal transduction of the proinflammatory cytokines IL-1 and TNF as well as of cellular stress in HMC.

Calcium-Calmodulin-Dependent Protein Kinases↗
PubMed1996

Surgery for biliary obstruction by tumour thrombus in primary liver cancer.

Five patients with primary liver cancer presented with obstructive jaundice due to extension of tumour thrombus into the biliary ducts. Three patients had hepatocellular carcinoma, two of whom had alcoholic cirrhosis, and the other two had a peripheral cholangiocarcinoma. Preoperative diagnosis of biliary thrombus was best achieved by ultrasonography and computed tomography which showed peripheral hepatic tumour with dilated bile ducts containing dense material. All patients underwent liver resection associated with biliary exploration, clearance and T-tube drainage. Major hepatectomy was required in four cases. There were no postoperative deaths; one patient developed a subphrenic collection of bile which was drained percutaneously. All patients survived more than a year; median survival was 29 months. There are two long-term survivors without recurrence at 29 and 80 months. Patients with primary liver cancer and jaundice due to migrated tumour fragments in the common bile duct may benefit from surgical resection which can result in long-term resolution of symptoms and occasional cure.

Carcinoma, Hepatocellular↗
PubMed1996

Interleukin-1 induced signalling: biphasic activation of mitogen-activated protein kinase kinase and mitogen-activated protein kinases in HeLa cells. Involvement of phosphoprotein phosphatases.

Results of this study document a biphasic activation of protein kinases of the MAP kinase cascade-MEK and MAP kinases-upon interleukin-1 stimulation in human HeLa cells. The specific activities of both MEK and MAP kinases were increased within 1 min, declined rapidly to control levels and increased again after 15 min of interleukin-1 stimulation. Inhibition by okadaic acid of serine/threonine specific phosphatases resulted in a marked increase in interleukin-1 stimulated MEK and MAP kinase activities. Elevation by interleukin-1 of the specific activities of MEK and MAP kinases correlated with suppression of serine/threonine phosphatases in the late phase of stimulation. The data indicate, that enhanced phosphorylation of cellular proteins by enzymes of the MAP kinase cascade might represent a fine balance between activated protein kinases and repressed phosphoprotein phosphatase 2A in interleukin-1 stimulated HeLa cells.

Calcium-Calmodulin-Dependent Protein Kinases↗