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)

Genetic and biochemical studies have identified kinase suppressor of Ras (KSR) to be a conserved component of Ras-dependent signaling pathways. To better understand the role of KSR in signal transduction, we have initiated studies investigating the effect of phosphorylation and protein interactions on KSR function. Here, we report the identification of five in vivo phosphorylation sites of KSR. In serum-starved cells, KSR contains two constitutive sites of phosphorylation (Ser297 and Ser392), which mediate the binding of KSR to the 14-3-3 family of proteins. In the presence of activated Ras, KSR contains three additional sites of phosphorylation (Thr260, Thr274, and Ser443), all of which match the consensus motif (Px[S/T]P) for phosphorylation by mitogen-activated protein kinase (MAPK). Further, we find that treatment of cells with the MEK inhibitor PD98059 blocks phosphorylation of the Ras-inducible sites and that activated MAPK associates with KSR in a Ras-dependent manner. Together, these findings indicate that KSR is an in vivo substrate of MAPK. Mutation of the identified phosphorylation sites did not alter the ability of KSR to facilitate Ras signaling in Xenopus oocytes, suggesting that phosphorylation at these sites may serve other functional roles, such as regulating catalytic activity. Interestingly, during the course of this study, we found that the biological effect of KSR varied dramatically with the level of KSR protein expressed. In Xenopus oocytes, KSR functioned as a positive regulator of Ras signaling when expressed at low levels, whereas at high levels of expression, KSR blocked Ras-dependent signal transduction. Likewise, overexpression of Drosophila KSR blocked R7 photoreceptor formation in the Drosophila eye. Therefore, the biological function of KSR as a positive effector of Ras-dependent signaling appears to be dependent on maintaining KSR protein expression at low or near-physiological levels.
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PMID:Identification of constitutive and ras-inducible phosphorylation sites of KSR: implications for 14-3-3 binding, mitogen-activated protein kinase binding, and KSR overexpression. 985 47

Genetic screens for modifiers of activated Ras phenotypes have identified a novel protein, kinase suppressor of Ras (KSR), which shares significant sequence homology with Raf family protein kinases. Studies using Drosophila melanogaster and Caenorhabditis elegans predict that KSR positively regulates Ras signaling; however, the function of mammalian KSR is not well understood. We show here that two predicted kinase-dead mutants of KSR retain the ability to complement ksr-1 loss-of-function alleles in C. elegans, suggesting that KSR may have physiological, kinase-independent functions. Furthermore, we observe that murine KSR forms a multimolecular signaling complex in human embryonic kidney 293T cells composed of HSP90, HSP70, HSP68, p50(CDC37), MEK1, MEK2, 14-3-3, and several other, unidentified proteins. Treatment of cells with geldanamycin, an inhibitor of HSP90, decreases the half-life of KSR, suggesting that HSPs may serve to stabilize KSR. Both nematode and mammalian KSRs are capable of binding to MEKs, and three-point mutants of KSR, corresponding to C. elegans loss-of-function alleles, are specifically compromised in MEK binding. KSR did not alter MEK activity or activation. However, KSR-MEK binding shifts the apparent molecular mass of MEK from 44 to >700 kDa, and this results in the appearance of MEK in membrane-associated fractions. Together, these results suggest that KSR may act as a scaffolding protein for the Ras-mitogen-activated protein kinase pathway.
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PMID:Kinase suppressor of Ras forms a multiprotein signaling complex and modulates MEK localization. 1040 42

In Drosophila melanogaster and Caenorhabditis elegans, kinase suppressor of Ras (KSR) functions as a positive modulator of Ras-dependent signaling either upstream of or parallel to Raf. Attempts to characterize the biochemical and biological properties of mammalian KSR, however, have had limited success. Although some studies demonstrated a requirement of KSR kinase activity for its action, others indicated the kinase function of KSR is dispensable and suggested that KSR acts primarily as a scaffold protein. Interpretations of KSR function are further hampered by the lack of a standardized assay for its kinase activity in vitro. To address this issue, we established a two-stage in vitro kinase assay in which KSR never comes in contact with any recombinant kinases other than c-Raf-1. Using this assay, we show that KSR immunoprecipitated from quiescent COS-7 cells overexpressing Flag-tagged KSR was inactive, but its activity was rapidly and markedly induced upon epidermal growth factor treatment. Moreover, KSR-reconstituted mitogen-activated protein kinase activation was detected in KSR immunoprecipitates depleted of all contaminating kinases (c-Raf-1, MEK1, ERK2) by multiple high salt washes. Only full-length kinase-active KSR was capable of signaling c-Raf-1-dependent activity as kinase inactive and C- and N-terminal deletion mutants were without effect. Furthermore, endogenous KSR isolated from A431 cells, which contain high levels of activated EGF receptor, displays constitutively enhanced kinase activity. Hence, KSR kinase activity is not an artifact of overexpression but a property intrinsic to this protein. The recognition of EGF as a potent activator of KSR kinase activity and the availability of a well defined in vitro kinase assay should facilitate the definition of the function of KSR as a Ras-effector molecule.
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PMID:Epidermal growth factor treatment enhances the kinase activity of kinase suppressor of Ras. 1076 33

Recent observations support the importance of ceramide synthesis de novo in the induction of apoptosis. However, the downstream targets of de novo-synthesized ceramide are unknown. Here we show that palmitate incorporated into ceramide and induced apoptotic DNA fragmentation in astrocytes. These effects of palmitate were exacerbated when fatty acid breakdown was uncoupled and were not evident in neurons, which show a very low capacity to take up and metabolize palmitate. Palmitate-induced apoptosis of astrocytes was prevented by L-cycloserine and fumonisin B1, two inhibitors of ceramide synthesis de novo, and by PD098059, an inhibitor of the extracellular signal-regulated kinase (ERK) cascade. Accordingly, palmitate activated ERK by a process that was dependent on ceramide synthesis de novo and Raf-1, but independent of kinase suppressor of Ras. Other potential targets of ceramide in the control of cell fate, namely, c-Jun amino-terminal kinase, p38 mitogen-activated protein kinase, and protein kinase B, were not significantly affected in astrocytes exposed to palmitate. Results show that the Raf-1/ERK cascade is the selective downstream target of de novo-synthesized ceramide in the induction of apoptosis in astrocytes and also highlight the importance of ceramide synthesis de novo in apoptosis of astrocytes, which might have pathophysiological relevance.
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PMID:De novo-synthesized ceramide signals apoptosis in astrocytes via extracellular signal-regulated kinase. 1105 53

kinase suppressor of Ras (ksr) encodes a putative protein kinase that by genetic criteria appears to function downstream of RAS in multiple receptor tyrosine kinase (RTK) pathways. While biochemical evidence suggests that the role of KSR is closely linked to the signal transduction mechanism of the MAPK cascade, the precise molecular function of KSR remains unresolved. To further elucidate the role of KSR and to identify proteins that may be required for KSR function, we conducted a dominant modifier screen in Drosophila based on a KSR-dependent phenotype. Overexpression of the KSR kinase domain in a subset of cells during Drosophila eye development blocks photoreceptor cell differentiation and results in the external roughening of the adult eye. Therefore, mutations in genes functioning with KSR might modify the KSR-dependent phenotype. We screened approximately 185,000 mutagenized progeny for dominant modifiers of the KSR-dependent rough eye phenotype. A total of 15 complementation groups of Enhancers and four complementation groups of Suppressors were derived. Ten of these complementation groups correspond to mutations in known components of the Ras1 pathway, demonstrating the ability of the screen to specifically identify loci critical for Ras1 signaling and further confirming a role for KSR in Ras1 signaling. In addition, we have identified 4 additional complementation groups. One of them corresponds to the kismet locus, which encodes a putative chromatin remodeling factor. The relevance of these loci with respect to the function of KSR and the Ras1 pathway in general is discussed.
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PMID:A genetic screen for modifiers of a kinase suppressor of Ras-dependent rough eye phenotype in Drosophila. 1106 97

Previous studies have demonstrated that a number of biochemical actions of ceramide are mediated through protein kinase signalling pathways, such as p42/p44 mitogen-activated protein kinase (p42/p44 MAPK) and c-Jun N-terminal directed protein kinase (JNK). Ceramide-activated protein kinases, such as the kinase suppressor of Ras (KSR) and protein kinase Czeta (PKCzeta), are involved in the regulation of c-Raf, which promotes sequential activation of MEK-1 and p42/p44 MAPK in mammalian cells. However, in cultured airway smooth muscle (ASM) cells, neither KSR nor PKCzeta are involved in the C2-ceramide (C2-Cer)-dependent activation of this kinase cascade. Instead, we found that C2-Cer utilises a novel pathway involving tyrosine kinases, phosphoinositide 3-kinase (PI3K) and conventional PKC isoform(s). We also found that despite its ability to stimulate p42/p44 MAPK, C2-Cer inhibited platelet-derived growth factor (PDGF)-stimulated DNA synthesis. The possibility that growth arrest could be mediated by JNK was discounted on the basis that PDGF, as well as ceramide, stimulated JNK in these cells. Therefore, growth arrest in response to ceramide is mediated by an alternative mechanism.
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PMID:Ceramide-dependent regulation of p42/p44 mitogen-activated protein kinase and c-Jun N-terminal-directed protein kinase in cultured airway smooth muscle cells. 1115 59

Mitogen-activated protein (MAP) kinase activity is essential for tumor necrosis factor (TNF) alpha receptor 1 regulation of intestinal epithelial cell proliferation. However, the mechanism of TNF-alpha mediated activation of extracellular signal-regulated kinase (ERK)/M1AP kinase has not been established clearly. Both TNF-alpha and cell-permeable ceramide have been reported to increase the kinase activity of kinase suppressor of Ras (KSR). To determine the role of KSR in TNF-alpha-induced ERK1/ERK2 activation, we studied young adult mouse colon cells expressing a dominant-negative, kinase-inactive (ki) KSR. We report that TNF-alpha, a cell-permeable ceramide, and sphingomyelinase stimulate ERK1/ERK2 activation and increase the phosphoserine content of KSR, which are inhibited by kiKSR expression in intact cells. Furthermore, TNF-alpha-induced Raf-1 threonine phosphorylation, kinase activity toward MEK1, and association with KSR are also inhibited by kiKSR expression. Our data also show by sequential in vitro kinase assays that TNF-alpha enhances KSR phosphorylation of Raf-1 on threonine, enhancing Raf-1 kinase activity toward MAP kinase kinase. We therefore conclude that KSR is an essential upstream regulator of TNF-alpha-stimulated ERK1/ERK2 activation, most likely mediated via direct phosphorylation of Raf-1.
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PMID:Kinase suppressor of ras is necessary for tumor necrosis factor alpha activation of extracellular signal-regulated kinase/mitogen-activated protein kinase in intestinal epithelial cells. 1122 91

The single layer of epithelial cells lining the intestine that serves as an important physical and functional barrier regulating the uptake of nutrients and the exclusion of various environmental antigens is disrupted in inflammatory bowel diseases. A central cytokine in the pathogenesis of inflammatory bowel disease is tumor necrosis factor (TNF), which increases apoptosis in a number of cell types. However, details determining the fate of intestinal cells exposed to high levels of TNF are lacking. Our laboratory reported that kinase suppressor of Ras (KSR) regulates TNF activation of the Raf/mitogen-activated protein (MAP) kinase/extracellular signal-regulated kinase (ERK) kinase/ERK signaling cassette by threonine phosphorylation of Raf-1, regulating proliferation and differentiation pathways. In the present study, we expressed a dominant-negative kinase-inactive KSR and determined the survival of young adult mouse colon cells exposed to TNF. Our data show that inhibition of KSR signaling decreases survival and increases apoptosis of TNF-treated cells. Antiapoptotic pathways including nuclear factor kappa B activation and one of its transcriptional targets, cIAP2 (c inhibitor of apoptosis protein 2) gene expression, and ERK/MAP kinase activation are all inhibited in TNF-treated kinase-inactive KSR-expressing young adult mouse colon cells. These antiapoptotic pathways are also inhibited by antisense-mediated down-regulation of KSR. However, TNF activation of p38 or stress-activated protein kinase/c-Jun NH(2)-terminal kinase is not inhibited by disruption of KSR signaling. Furthermore, inhibitors of both ERK and nuclear factor kappa B activation synergistically enhance apoptosis of cells treated with TNF. These findings demonstrate that KSR plays a novel regulatory role in intestinal epithelial cells exposed to TNF by activating cell survival pathways.
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PMID:Kinase suppressor of Ras determines survival of intestinal epithelial cells exposed to tumor necrosis factor. 1175 83

While scaffold proteins are thought to be key components of signaling pathways, their exact function is unknown. By preassembling multiple components of signaling cascades, scaffolds are predicted to influence the efficiency and/or specificity of signaling events. Here we analyze a potential scaffold of the Ras/mitogen-activated protein kinase (MAPK) pathway, kinase suppressor of Ras (KSR), by generating KSR-deficient mice. KSR-deficient mice were grossly normal even though ERK kinase activation was attenuated to a degree sufficient to block T-cell activation and inhibit tumor development. Consistent with its role as a scaffold, high-molecular-weight complexes containing KSR, MEK, and ERK were lost in the absence of KSR. This demonstrates that KSR is a bona fide scaffold that is not required for but enhances signaling via the Ras/MAPK signaling pathway.
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PMID:Kinase suppressor of Ras (KSR) is a scaffold which facilitates mitogen-activated protein kinase activation in vivo. 1194 Jun 61

Raabe and Rapp discuss how the scaffolding proteins may spatially localize mitogen-activated protein kinase (MAPK) signaling cassettes to allow signaling specificity. They focus on the adaptor kinase suppressor of Ras (KSR) and describe how the interactions between KSR and other proteins and the subcellular localization of KSR are regulated by phosphorylation.
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PMID:KSR--a regulator and scaffold protein of the MAPK pathway. 1206 Jul 87


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