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MAPK Cascades (mapk + cascade)
Selected AbstractsRegulation of mitogen-activated protein kinases by glutamate receptorsJOURNAL OF NEUROCHEMISTRY, Issue 1 2007John Q. Wang Abstract Glutamate receptors regulate gene expression in neurons by activating intracellular signaling cascades that phosphorylate transcription factors within the nucleus. The mitogen-activated protein kinase (MAPK) cascade is one of the best characterized cascades in this regulatory process. The Ca2+ -permeable ionotropic glutamate receptor, mainly the NMDA receptor subtype, activates MAPKs through a biochemical route involving the Ca2+ -sensitive Ras-guanine nucleotide releasing factor, Ca2+/calmodulin-dependent protein kinase II, and phosphoinositide 3-kinase. The metabotropic glutamate receptor (mGluR), however, activates MAPKs primarily through a Ca2+ -insensitve pathway involving the transactivation of receptor tyrosine kinases. The adaptor protein Homer also plays a role in this process. As an information superhighway between surface glutamate receptors and transcription factors in the nucleus, active MAPKs phosphorylate specific transcription factors (Elk-1 and CREB), and thereby regulate distinct programs of gene expression. The regulated gene expression contributes to the development of multiple forms of synaptic plasticity related to long-lasting changes in memory function and addictive properties of drugs of abuse. This review, by focusing on new data from recent years, discusses the signaling mechanisms by which different types of glutamate receptors activate MAPKs, features of each MAPK cascade in regulating gene expression, and the importance of glutamate/MAPK-dependent synaptic plasticity in memory and addiction. [source] The neuronal MAP kinase cascade: a biochemical signal integration system subserving synaptic plasticity and memoryJOURNAL OF NEUROCHEMISTRY, Issue 1 2001J. David Sweatt The mitogen-activated protein kinase (MAP kinase, MAPK) cascade, as the name implies, was originally discovered as a critical regulator of cell division and differentiation. As further details of this signaling cascade were worked out, it became clear that the MAPK cascade is in fact a prototype for a family of signaling cascades that share the motif of three serially linked kinases regulating each other by sequential phosphorylation. Thus, a revised nomenclature arose that uses the term MAPK to refer to the entire superfamily of signaling cascades (comprising the erks, the JNKs and the p38 stress activated protein kinases), and specifies the prototype MAPK as the extracellular signal-regulated kinase (erk). The two erk MAPK isoforms, p44 MAPK and p42 MAPK, are referred to as erk1 and erk2, respectively. The erks are abundantly expressed in neurons in the mature central nervous system, raising the question of why the prototype molecular regulators of cell division and differentiation are present in these non-dividing, terminally differentiated neurons. This review will describe the beginnings of an answer to this question. Interestingly, the general model has begun to emerge that the erk signaling system has been co-opted in mature neurons to function in synaptic plasticity and memory. Moreover, recent insights have led to the intriguing prospect that these molecules serve as biochemical signal integrators and molecular coincidence detectors for coordinating responses to extracellular signals in neurons. In this review I will first outline the essential components of this signal transduction cascade, and briefly describe recent results implicating the erks in mammalian synaptic plasticity and learning. I will then proceed to outline recent results implicating the erks as molecular signal integrators and, potentially, coincidence detectors. Finally, I will speculate on what the critical downstream effectors of the erks are in neurons, and how they might provide a readout of the integrated signal. [source] Fungicide activity through activation of a fungal signalling pathwayMOLECULAR MICROBIOLOGY, Issue 6 2004Kaihei Kojima Summary Fungicides generally inhibit enzymatic reactions involved in fungal cellular biosynthesis. Here we report, for the first time, an example of fungicidal effects through hyperactivation of a fungal signal transduction pathway. The OSC1 gene, encoding a MAP kinase (MAPK) related to yeast Hog1, was isolated from the fungal pathogen Colletotrichum lagenarium that causes cucumber anthracnose. The osc1 knockout mutants were sensitive to high osmotic stress and showed increased resistance to the fungicide fludioxonil, indicating that Osc1 is involved in responses to hyperosmotic stress and sensitivity to fludioxonil. The Osc1 MAPK is phosphorylated under high osmotic conditions, indicating activation of Osc1 by high osmotic stress. Importantly, fludioxonil treatment also activates phosphorylation of Osc1, suggesting that improper activation of Osc1 by fludioxonil has negative effects on fungal growth. In the presence of fludioxonil, the wild-type fungus was not able to infect the host plant because of a failure of appressorium-mediated penetration, whereas osc1 mutants successfully infected plants. Analysis using a OSC1- GFP fusion gene indicated that Osc1 is rapidly translocated to the nucleus in appressorial cells after the addition of fludioxonil, suggesting that fludioxonil impairs the function of infection structures by activation of Osc1. Furthermore, fludioxonil activates Hog1-type MAPKs in the plant pathogenic fungi Cochliobolus heterostrophus and Botrytis cinerea. These results strongly suggest that fludioxonil acts as a fungicide, in part, through activation of the MAPK cascade in fungal pathogens. [source] Mss11p is a transcription factor regulating pseudohyphal differentiation, invasive growth and starch metabolism in Saccharomyces cerevisiae in response to nutrient availabilityMOLECULAR MICROBIOLOGY, Issue 1 2003Marco Gagiano Summary In Saccharomyces cerevisiae, the cell surface protein, Muc1p, was shown to be critical for invasive growth and pseudohyphal differentiation. The transcription of MUC1 and of the co-regulated STA2 glucoamylase gene is controlled by the interplay of a multitude of regulators, including Ste12p, Tec1p, Flo8p, Msn1p and Mss11p. Genetic analysis suggests that Mss11p plays an essential role in this regulatory process and that it functions at the convergence of at least two signalling cascades, the filamentous growth MAPK cascade and the cAMP-PKA pathway. Despite this central role in the control of filamentous growth and starch metabolism, the exact molecular function of Mss11p is unknown. We subjected Mss11p to a detailed molecular analysis and report here on its role in transcriptional regulation, as well as on the identification of specific domains required to confer transcriptional activation in response to nutritional signals. We show that Mss11p contains two independent transactivation domains, one of which is a highly conserved sequence that is found in several proteins with unidentified function in mammalian and invertebrate organisms. We also identify conserved amino acids that are required for the activation function. [source] Identification of a putative voltage-gated Ca2+ channel as a key regulator of elicitor-induced hypersensitive cell death and mitogen-activated protein kinase activation in riceTHE PLANT JOURNAL, Issue 6 2005Takamitsu Kurusu Summary Elicitor-triggered transient membrane potential changes and Ca2+ influx through the plasma membrane are thought to be important during defense signaling in plants. However, the molecular bases for the Ca2+ influx and its regulation remain largely unknown. Here we tested effects of overexpression as well as retrotransposon (Tos17)-insertional mutagenesis of the rice two-pore channel 1 (OsTPC1), a putative voltage-gated Ca2+ -permeable channel, on a proteinaceous fungal elicitor-induced defense responses in rice cells. The overexpressor showed enhanced sensitivity to the elicitor to induce oxidative burst, activation of a mitogen-activated protein kinase (MAPK), OsMPK2, as well as hypersensitive cell death. On the contrary, a series of defense responses including the cell death and activation of the MAPK were severely suppressed in the insertional mutant, which was complemented by overexpression of the wild-type gene. These results suggest that the putative Ca2+ -permeable channel determines sensitivity to the elicitor and plays a role as a key regulator of elicitor-induced defense responses, activation of MAPK cascade and hypersensitive cell death. [source] Function of a mitogen-activated protein kinase pathway in N gene-mediated resistance in tobaccoTHE PLANT JOURNAL, Issue 4 2003Hailing Jin Summary The active defense of plants against pathogens often includes rapid and localized cell death known as hypersensitive response (HR). Protein phosphorylation and dephosphorylation are implicated in this event based on studies using protein kinase and phosphatase inhibitors. Recent transient gain-of-function studies demonstrated that the activation of salicylic acid-induced protein kinase (SIPK) and wounding-induced protein kinase (WIPK), two tobacco mitogen-activated protein kinases (MAPKs) by their upstream MAPK kinase (MAPKK), NtMEK2 leads to HR-like cell death. Here, we report that the conserved kinase interaction motif (KIM) in MAPKKs is required for NtMEK2 function. Mutation of the conserved basic amino acids in this motif, or the deletion of N-terminal 64 amino acids containing this motif significantly compromised or abolished the ability of NtMEK2DD to activate SIPK/WIPK in vivo. These mutants were also defective in interacting with SIPK and WIPK, suggesting protein,protein interaction is required for the functional integrity of this MAPK cascade. To eliminate Agrobacterium that is known to activate a number of defense responses in transient transformation experiments, we generated permanent transgenic plants. Induction of NtMEK2DD expression by dexamethasone induced HR-like cell death in both T1 and T2 plants. In addition, by using PVX-induced gene silencing, we demonstrated that the suppression of all three known components in the NtMEK2,SIPK/WIPK pathway attenuated N gene-mediated TMV resistance. Together with previous report that SIPK and WIPK are activated by TMV in a gene-for-gene-dependent manner, we conclude that NtMEK2,SIPK/WIPK pathway plays a positive role in N gene-mediated resistance, possibly through regulating HR cell death. [source] Constitutive activation of MAPK cascade in acute quadriplegic myopathyANNALS OF NEUROLOGY, Issue 2 2004Simone Di Giovanni MD Acute quadriplegic myopathy (AQM; also called "critical illness myopathy") shows acute muscle wasting and weakness and is experienced by some patients with severe systemic illness, often associated with administration of corticosteroids and/or neuroblocking agents. Key aspects of AQM include muscle atrophy and myofilament loss. Although these features are shared with neurogenic atrophy, myogenic atrophy in AQM appears mechanistically distinct from neurogenic atrophy. Using muscle biopsies from AQM, neurogenic atrophy, and normal controls, we show that both myogenic and neurogenic atrophy share induction of myofiber-specific ubiquitin/proteosome pathways (eg, atrogin-1). However, AQM patient muscle showed a specific strong induction of transforming growth factor (TGF),,/MAPK pathways. Atrophic AQM myofibers showed coexpression of TGF-, receptors, p38 MAPK, c-jun, and c-myc, including phosphorylated active forms, and these same fibers showed apoptotic features. Our data suggest a model of AQM pathogenesis in which stress stimuli (sepsis, corticosteroids, pH imbalance, osmotic imbalance) converge on the TGF-, pathway in myofibers. The acute stimulation of the TGF-,/MAPK pathway, coupled with the inactivity-induced atrogin-1/proteosome pathway, leads to the acute muscle loss seen in AQM patients. Ann Neurol 2004 [source] Mitogen-activated protein kinase signal transduction in skeletal muscle: effects of exercise and muscle contractionACTA PHYSIOLOGICA, Issue 3 2001U. Widegren 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. [source] Oncostatin M enhances the expression of prostaglandin E2 and cyclooxygenase-2 in astrocytes: Synergy with interleukin-1,, tumor necrosis factor-,, and bacterial lipopolysaccharideGLIA, Issue 4 2003Pavle Repovic Abstract Oncostatin M (OSM), a cytokine of the interleukin-6 family, is expressed in rheumatoid arthritis, multiple sclerosis, multiple myeloma, and other inflammatory and neoplastic conditions. Prostaglandin E2 (PGE2), an eicosanoid also associated with inflammation and cancer, has recently been shown to induce OSM expression. We report here that OSM in turn induces PGE2 production by astrocytes and astroglioma cells. More importantly, in combination with the inflammatory mediators IL-1,, tumor necrosis factor-,, and lipopolysaccharide, OSM exhibits a striking synergy, resulting in up to 50-fold higher PGE2 production by astrocytes, astroglioma, and neuroblastoma cell lines. Enhanced PGE2 production by OSM and IL-1, treatment is explained by their effect on cyclooxygenase-2 (COX-2), an enzyme that catalyzes the committed step in PGE2 synthesis. Of the enzymes involved in PGE2 biosynthesis, only COX-2 mRNA and protein levels are synergistically amplified by OSM and IL-1,. Nuclear run-on assays demonstrate that OSM and IL-1, synergistically upregulate transcription of the COX-2 gene, and the mRNA stability assay indicates that COX-2 mRNA is posttranscriptionally stabilized by OSM and IL-1,. To effect synergy on the PGE2 level, OSM signals in part through its gp130/OSMR, receptor, since neutralizing antibodies against gp130 and OSMR,, but not LIFR,, decrease PGE2 production in response to OSM plus IL-1,. SB202190 and U0126, inhibitors of p38 MAPK and ERK1/2 activation, respectively, inhibit IL-1, and OSM upregulation of COX-2 and PGE2, indicating that these MAPK cascades are utilized by both stimuli. This mechanism of PGE2 amplification may be active in brain pathologies where both OSM and IL-1, are present, such as glioblastomas and multiple sclerosis. GLIA 42:433,446, 2003. © 2003 Wiley-Liss, Inc. [source] Osteoblast-Derived TGF-,1 Stimulates IL-8 Release Through AP-1 and NF-,B in Human Cancer Cells,JOURNAL OF BONE AND MINERAL RESEARCH, Issue 6 2008Yi-Chin Fong Abstract Introduction: The bone marrow microenvironment is further enriched by growth factors released during osteoclastic bone resorption. It has been reported that the chemokine interleukin (IL)-8 is a potent and direct activator of osteoclastic differentiation and bone resorption. However, the effect of bone-derived growth factors on the IL-8 production in human cancer cells and the promotion of osteoclastogenesis are largely unknown. The aim of this study was to investigate whether osteoblast-derived TGF-,1 is associated with osteolytic bone diseases. Materials and Methods: IL-8 mRNA levels were measured using RT-PCR analysis. MAPK phosphorylation was examined using the Western blot method. siRNA was used to inhibit the expression of TGF-,1, BMP-2, and IGF-1. DNA affinity protein-binding assay and chromatin immunoprecipitation assays were used to study in vitro and in vivo binding of c- fos, c- jun, p65, and p50 to the IL-8 promoter. A transient transfection protocol was used to examine IL-8, NF-,B, and activator protein (AP)-1 activity. Results: Osteoblast conditioned medium (OBCM) induced activation of IL-8, AP-1, and NF-,B promoter in human cancer cells. Osteoblasts were transfected with TGF-,1, BMP-2, or IGF-1 small interfering RNA, and the medium was collected after 48 h. TGF-,1 but not BMP-2 or IGF-1 siRNA inhibited OBCM-induced IL-8 release in human cancer cells. In addition, TGF-,1 also directly induced IL-8 release in human cancer cells. Activation of AP-1 and NF-,B DNA-protein binding and MAPKs after TGF-,1 treatment was shown, and TGF-,1,induced IL-8 promoter activity was inhibited by the specific inhibitors of MAPK cascades. Conclusions: In this study, we provide evidence to show that the osteoblasts release growth factors, including TGF-,1, BMP-2, and IGF-1. TGF-,1 is the major contributor to the activation of extracellular signal-related kinase (ERK), p38, and c-Jun N-terminal kinase (JNK), leading to the activation of AP-1 and NF-,B on the IL-8 promoter and initiation of IL-8 mRNA and protein release, thereby promoting osteoclastogenesis. [source] Bradykinin-induced p42/p44 MAPK phosphorylation and cell proliferation via Src, EGF receptors, and PI3-K/Akt in vascular smooth muscle cellsJOURNAL OF CELLULAR PHYSIOLOGY, Issue 3 2005Chuen-Mao Yang In our previous study, bradykinin (BK) exerts its mitogenic effect through Ras/Raf/MEK/MAPK pathway in vascular smooth muscle cells (VSMCs). In addition to this pathway, the non-receptor tyrosine kinases (Src), EGF receptor (EGFR), and phosphatidylinositol 3-kinase (PI3-K) have been implicated in linking a variety of G-protein coupled receptors to MAPK cascades. Here, we investigated whether these different mechanisms participating in BK-induced activation of p42/p44 MAPK and cell proliferation in VSMCs. We initially observed that BK- and EGF-dependent activation of Src, EGFR, Akt, and p42/p44 MAPK and [3H]thymidine incorporation were mediated by Src and EGFR, because the Src inhibitor PP1 and EGFR kinase inhibitor AG1478 abrogated BK- and EGF-dependent effects. Inhibition of PI3-K by LY294002 attenuated BK-induced Akt and p42/p44 MAPK phosphorylation and [3H]thymidine incorporation, but had no effect on EGFR phosphorylation, suggesting that EGFR may be an upstream component of PI3-K/Akt and MAPK in these responses. This hypothesis was supported by the tranfection with dominant negative plasmids of p85 and Akt which significantly attenuated BK-induced Akt and p42/p44 MAPK phosphorylation. Pretreatment with U0126 (a MEK1/2 inhibitor) attenuated the p42/p44 MAPK phosphorylation and [3H]thymidine incorporation stimulated by BK, but had no effect on Akt activation. Moreover, BK-induced transactivation of EGFR and cell proliferation was blocked by matrix metalloproteinase inhibitor GM6001. These results suggest that, in VSMCs, the mechanism of BK-stimulated activation of p42/p44 MAPK and cell proliferation was mediated, at least in part, through activation of Src family kinases, EGFR transactivation, and PI3-K/Akt. Copyright © 2004 Wiley-Liss, Inc. [source] AUF-1 mediates inhibition by nitric oxide of lipopolysaccharide-induced matrix metalloproteinase-9 expression in cultured astrocytesJOURNAL OF NEUROSCIENCE RESEARCH, Issue 2 2006Wenlan Liu Abstract Neuroinflammatory diseases are associated with increased production of matrix metalloproteinase-9 (MMP-9) and excessive generation of nitric oxide (NO). NO hasbeen reported to have variable effects on MMP-9 gene expression and activation in various cell types. Inthe present study, we investigated the effect of NOon MMP-9 expression in primary cortical astrocytes. Zymography and real-time PCR showed that lipopolysaccharide (LPS) dramatically increased latent MMP-9 gelatinolytic activity and MMP-9 mRNA expression. By using the NO donor DETA NONOate, we observed a dose-dependent inhibition of MMP-9 induction by LPS. Active forms of MMP-9 were not found by zymography after NO treatment. The MEK1/2 inhibitor U0126 completely inhibited LPS-induced MMP-9, which was partially inhibited by the p38 MAPK inhibitor SB203580. NO had no effect on LPS-stimulated ERK1/2 and p38 MAPK activation, suggesting that the inhibitory action of NO occurs downstream of MAPK cascades. Real-time PCR analysis showed that NO accelerated the degradation of MMP-9 mRNA after LPS induction. Western blotting and pull-down assay demonstrated that NO increased AUF-1 expression as well as its specific binding to the MMP-9 gene 3,-untranslated region. Knockdown of AUF-1 with siRNA partially reversed the inhibitory action of NO on LPS-stimulated MMP-9 induction. We conclude that NO does not activate MMP-9 in astrocyte cultures but reduces LPS-induced MMP-9 expression via accelerating MMP-9 mRNA degradation, which is partially mediated by AUF-1. Our results suggest that elevated NO concentrations may suppress MMP-9 and restrict the inflammatory response in neurodegenerative diseases. © 2006 Wiley-Liss, Inc. [source] Ras-mediated intestinal epithelial cell transformation requires cyclooxygenase-2-induced prostaglandin E2 signalingMOLECULAR CARCINOGENESIS, Issue 12 2007Gretchen A. Repasky Abstract Ras-mediated transformation is associated with upregulation of cyclooxygenase-2 (COX-2), which in turn promotes prostaglandin E2 (PGE2) synthesis and secretion. Although recent studies have identified molecular mechanisms by which Ras mediates upregulation of COX-2, conflicting observations have been made. Furthermore, while COX-2 upregulation has been shown to be important for Ras transformation, the signaling pathways initiated by PGE2 -stimulation of EP family of heterotrimeric G protein-coupled receptors (GPCR) and contribution of PGE2 signaling to Ras-mediated transformation are issues that remain unresolved. In this study, we first determined that Raf effector pathway activation of the extracellular-regulated kinase (ERK) mitogen-activated protein kinase (MAPK) cascade alone was sufficient and necessary for COX-2 and PGE2 upregulation. However, Raf-independent regulation of the c- jun N-terminal kinase (JNK) and p38 MAPK cascades is also involved in COX-2 and PGE2 upregulation, with the JNK and p38 pathways exhibiting opposing roles in COX-2 and PGE2 upregulation. Furthermore, in contrast to previous studies, we found that an epidermal growth factor (EGF) receptor autocrine growth mechanism, another Raf-independent signaling mechanism, was necessary for COX-2 and PGE2 upregulation. Second, we determined that inhibition of EP1/2 receptor function blocked growth transformation by Ras, demonstrating that PGE2 upregulation is a key transforming function of COX-2. Finally, we found that PGE2 stimulated the activation of Ras and ERK, but not Akt, and reduced matrix deprivation-induced apoptosis, in untransformed epithelial cells. In summary, our studies define additional, multiple signaling mechanisms that promote COX-2 and PGE2 expression and show that COX-2-stimulated PGE2 -EP receptor signaling is required for growth and survival transformation by Ras. © 2007 Wiley-Liss, Inc. [source] Ultraviolet radiation stimulates expression of Snail family transcription factors in keratinocytesMOLECULAR CARCINOGENESIS, Issue 4 2007Laurie G. Hudson Abstract The related zinc finger transcription factors Slug and Snail modulate epithelial mesenchymal transformation (EMT), the conversion of sessile epithelial cells into migratory fibroblast-like cells. EMT occurs during development, wound healing, and tumor progression. Growth factors, acting through mitogen-activated protein kinase (MAPK) cascades, regulate expression of Slug and Snail. Expression of Snail family transcription factors appears to be elevated in UVR-induced murine squamous cell carcinomas (SCC). We report here that ultraviolet radiation (UVR), which activates MAPK cascades, also stimulates Snail and Slug expression in epidermal keratinocytes. UVR exposure transiently elevated Slug and Snail mRNA expression in human keratinocytes in vitro and mouse epidermis in vivo. This induction was mediated, at least in part, through the ERK and p38 MAPK cascades, as pharmacological inhibition of these cascades partially or completely blocked Slug and Snail induction by UVR. On the other hand, UVR induction of Slug and Snail was enhanced by inhibition of JNK. Slug appears to play a functional role in the acute response of keratinocytes to UVR, as UVR induction of keratin 6 in the epidermis of Slug knockout mice was markedly delayed compared to wild-type mice. Slug and Snail are known to regulate molecules important in the cytoskeleton, intercellular adhesion, cell motility, and apoptosis, thus it seems probable that transiently or persistently elevated expression of these factors fosters the progression of UVR-induced SCC. © 2007 Wiley-Liss, Inc. [source] Copper treatment activates mitogen-activated protein kinase signalling in ricePHYSIOLOGIA PLANTARUM, Issue 3 2003Chuan-Ming Yeh It is well known that mitogen-activated protein kinase (MAPK) pathways are modules involved in the transduction of extracellular signals to intracellular targets in all eukaryotes. In plants, it has been shown that MAPKs play a role in the signalling of biotic and abiotic stresses. To characterize signalling pathways involved in heavy metal-induced stress responses, we examine whether plant MAPKs are also involved in this process. The analyses of mRNA levels of OsMAPK genes have shown that only OsMAPK2 mRNA transcripts increased within 12 h upon CuCl2 treatment in suspension cells and roots. An in-gel kinase assay revealed that three protein kinases, approximate 42, 50, and 64-kDa, were activated by CuCl2 treatments. The approximate 42-kDa protein kinase displayed MAPK properties. Antioxidant, GSH, prevented copper-induced kinase activity. Furthermore, we found that rice roots underwent a rapid cell death upon this copper treatment. The copper-induced cell death of rice roots was partially blocked by MAPK kinase inhibitor, PD98059. These results suggest that the MAPK cascades may function in the plant heavy metal induced-signalling pathway. [source] Polymerization of the SAM domain of MAPKKK Ste11 from the budding yeast: Implications for efficient signaling through the MAPK cascadesPROTEIN SCIENCE, Issue 3 2005Surajit Bhattacharjya Abstract The sterile ,-motif (SAM) is a protein module ,70 residues long and mainly involved in the protein,protein interactions of cell signaling and transcriptional repression. The SAM domain of the yeast MAPKKK Ste11 has a well-folded dimeric structure in solution. Interestingly, the well-folded dimer of the Ste11 SAM undergoes a time-dependent self-assembly upon lowering of the pH, leading to the formation of high molecular weight oligomers. The oligomeric structures rapidly disassemble to the well-folded dimer upon reversal of the pH to close to neutral conditions. Circular dichroism (CD) and atomic force microscopy (AFM) experiments demonstrate that the oligomeric structure formed at pH 5.0 appears to be highly helical and has architecture akin to proto-fibrils. Residue-specific kinetics of pH-triggered oligomerization obtained from real-time 15N- 1H HSQC experiments indicate that the dimer-oligomer transition appears to involve all residues of the well-folded dimeric structure of the Ste11 SAM. Very interestingly, the interactions of the Ste11 and Ste50 SAM domains also lead to the formation of non-homogeneous hetero-complexes with significant populations of high molecular weight aggregates. AFM imaging shows that the Ste11-Ste50 hetero-polymeric aggregates assume the shapes of circular nano-particles with dimensions of 50,60 nano-meters (nm), in contrast to the proto-fibrils formed by the Ste11 SAM domain alone. Such intrinsic propensity for dimer to oligomer transition of the Ste50-binding SAM domain of Ste11 may endow the MAPKKK Ste11 with unique functional properties required for efficient and high fidelity signal transduction in the budding yeast. [source] Arabidopsis mitogen-activated protein kinase MPK12 interacts with the MAPK phosphatase IBR5 and regulates auxin signalingTHE PLANT JOURNAL, Issue 6 2009Jin Suk Lee Summary Mitogen-activated protein kinase (MAPK) phosphatases are important negative regulators in the MAPK signaling pathways responsible for many essential processes in plants, including development, stress management and hormonal responses. A mutation in INDOLE-3-BUTYRIC ACID-RESPONSE5 (IBR5), which is predicted to encode a dual-specificity MAPK phosphatase, was previously reported to confer reduced sensitivity to auxin and ABA in Arabidopsis roots. To further characterize IBR5, and to understand how it might help integrate MAPK cascades with hormone signaling, we searched for IBR5-interacting MAPKs. Yeast two-hybrid assays, in vitro binding assays and in vivo protein co-immunoprecipitation studies demonstrated that MPK12 and IBR5 are physically coupled. The C-terminus of MPK12 appears to be essential for its interaction with IBR5, and in vitro dephosphorylation and immunocomplex kinase assays indicated that activated MPK12 is efficiently dephosphorylated and inactivated by IBR5. MPK12 and IBR5 mRNAs are both widely expressed across Arabidopsis tissues, and at the subcellular level each protein is predominantly localized in the nucleus. In transgenic plants with reduced expression of the MPK12 gene, root growth is hypersensitive to exogenous auxins, but shows normal ABA sensitivity. MPK12 suppression in an ibr5 background partially complements the ibr5 auxin-insensitivity phenotype. Our results demonstrate that IBR5 is a bona fide MAPK phosphatase, and suggest that MPK12 is both a physiological substrate of IBR5 and a novel negative regulator of auxin signaling in Arabidopsis. [source] | ||||||||||||||||||||||