Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.1.3.16 (calcineurin)
 17,112 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Mitogen-activated protein (MAP) kinase phosphatase-1 (MKP-1) is the archetypal member of the dual-specificity protein phosphatase family, the expression of which can be rapidly induced by a variety of growth factors and cellular stress. Since MKP-1 protein localizes in the nucleus, it has been suggested to play an important role in the feedback control of MAP kinase-regulated gene transcription. Recently it has been demonstrated that the interaction of several cytosolic MAP kinase phosphatases with MAP kinases can trigger the catalytic activation of the phosphatases. It is unclear whether such a regulatory mechanism can apply to nuclear MAP kinase phosphatases and serve as an additional apparatus for the feedback control of MAP kinase-mediated gene expression. Here we have shown that MKP-1 associates directly with p38 MAP kinase both in vivo and in vitro, and that this interaction enhances the catalytic activity of MKP-1. The point mutation Asp-316-->Asn in the C-terminus of p38, analogous to the ERK2 (extracellular-signal-regulated kinase 2) sevenmaker mutation, dramatically decreases its binding to MKP-1 and substantially compromises its stimulatory effect on the catalytic activity of this phosphatase. Consistent with its defective interaction with MKP-1, this p38 mutant also displays greater resistance to dephosphorylation by the phosphatase. Our studies provide the first example of catalytic activation of a nuclear MAP kinase phosphatase through direct binding to a MAP kinase, suggesting that such a regulatory mechanism may play an important role in the feedback control of MAP kinase signalling in the nuclear compartment.
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PMID:Catalytic activation of mitogen-activated protein (MAP) kinase phosphatase-1 by binding to p38 MAP kinase: critical role of the p38 C-terminal domain in its negative regulation. 1106 68

Conventional qualitative approaches to signal transduction provide powerful ways to explore the architecture and function of signaling pathways. However, at the level of the complete system, they do not fully depict the interactions between signaling and metabolic pathways and fail to give a manageable overview of the complexity that is often a feature of cellular signal transduction. Here, we introduce a quantitative experimental approach to signal transduction that helps to overcome these difficulties. We present a quantitative analysis of signal transduction during early mitogen stimulation of lymphocytes, with steady-state respiration rate as a convenient marker of metabolic stimulation. First, by inhibiting various key signaling pathways, we measure their relative importance in regulating respiration. About 80% of the input signal is conveyed via identifiable routes: 50% through pathways sensitive to inhibitors of protein kinase C and MAP kinase and 30% through pathways sensitive to an inhibitor of calcineurin. Second, we quantify how each of these pathways differentially stimulates functional units of reactions that produce and consume a key intermediate in respiration: the mitochondrial membrane potential. Both the PKC and calcineurin routes stimulate consumption more strongly than production, whereas the unidentified signaling routes stimulate production more than consumption, leading to no change in membrane potential despite increased respiration rate. The approach allows a quantitative description of the relative importance of signal transduction pathways and the routes by which they activate a specific cellular process. It should be widely applicable.
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PMID:Quantitation of signal transduction. 1109 77

Insulin receptor-substrate-1 (IRS-1) is a docking protein for several tyrosine kinase receptors. Upon tyrosine phosphorylation, IRS-1 binds to signaling molecules that express Src homology 2 (SH-2) binding domains, including phosphatidylinositol 3-kinase (PI 3-kinase), phosphotyrosine phosphatase SHP-2 (Syp), Nck, Crk and Grb-2. Hydrogen peroxide (H(2)O(2)) induces tyrosine phosphorylation of key signaling mediators presumably by inhibition of tyrosine phosphatases. In many cell types, the activation of extracellular signal-related kinases (e.g. MAPK) and other protein kinases by H(2)O(2) leads to transcriptional activation. In the current study, we examined the effect of H(2)O(2) on IRS-1 tyrosine phosphorylation in primary cultured rat cerebellar granule neurons. H(2)O(2) stimulated the rapid tyrosine phosphorylation of IRS-1 and p42/p44 MAP kinase, and induced its association with PI 3-kinase. H(2)O(2)-induced IRS-1 phosphorylation was rapidly reversible (5 min) whereas MAPK phosphorylation persisted for up to 1 h. NMDA reversed H(2)O(2)-mediated tyrosine phosphorylation of IRS-1 and its association with PI 3-kinase. The dephosphorylation of IRS-1 by NMDA was calcium-dependent and was inhibited by the calcineurin inhibitor cyclosporine. Calmodulin-dependent tyrosine phosphatase activity of calcineurin was observed in vitro using both immunoprecipitated and recombinant tyrosine-phosphorylated IRS-1 as substrates. These data highlight the role of multiple phosphatases in the regulation of IRS-1 tyrosine phosphorylation and identify a novel functional property of calcineurin.
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PMID:Tyrosine phosphorylation of insulin receptor substrate-1 (IRS-1) by oxidant stress in cerebellar granule neurons: modulation by N-methyl-D-aspartate through calcineurin activity. 1127 62

Opitz syndrome (OS) is a human genetic disease characterized by deformities such as cleft palate that are attributable to defects in embryonic development at the midline. Gene mapping has identified OS mutations within a protein called Mid1. Wild-type Mid1 predominantly colocalizes with microtubules, in contrast to mutant versions of Mid1 that appear clustered in the cytosol. Using yeast two-hybrid screening, we found that the alpha4-subunit of protein phosphatases 2A/4/6 binds Mid1. Epitope-tagged alpha4 coimmunoprecipitated endogenous or coexpressed Mid1 from COS7 cells, and this required only the conserved C-terminal region of alpha4. Localization of Mid1 and alpha4 was influenced by one another in transiently transfected cells. Mid1 could recruit alpha4 onto microtubules, and high levels of alpha4 could displace Mid1 into the cytosol. Metabolic (32)P labeling of cells showed that Mid1 is a phosphoprotein, and coexpression of full-length alpha4 decreased Mid1 phosphorylation, indicative of a functional interaction. Association of green fluorescent protein-Mid1 with microtubules in living cells was perturbed by inhibitors of MAP kinase activation. The conclusion is that Mid1 association with microtubules, which seems important for normal midline development, is regulated by dynamic phosphorylation involving MAP kinase and protein phosphatase that is targeted specifically to Mid1 by alpha4. Human birth defects may result from environmental or genetic disruption of this regulatory cycle.
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PMID:Phosphorylation and microtubule association of the Opitz syndrome protein mid-1 is regulated by protein phosphatase 2A via binding to the regulatory subunit alpha 4. 1137 18

Long-term facilitation at the crayfish opener muscle is elicited by prolonged high frequency stimulation, and arises from an increase in functional active zones, resulting in increased transmitter release. LTF induction depends critically upon presynaptic calcium accumulation and calcineurin (PP2B) activity. The protein synthesis dependence of this synaptic strengthening was investigated. LTF occurred without transcription, but the translation inhibitors cycloheximide and anisomycin, or local presynaptic injection of mRNA cap analog m7GpppG, impaired LTF expression. Both MAP kinase and phosphatidylinositol 3-OH kinase (PI3K) activation are implicated in this rapamycin-sensitive synaptic potentiation. This study defines an important role for protein synthesis in the expression of activity-dependent plasticity, and provides mechanistic insight for the induction of this process at presynaptic sites.
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PMID:Phosphorylation and local presynaptic protein synthesis in calcium- and calcineurin-dependent induction of crayfish long-term facilitation. 1170 59

Phosphorylation of yeast 6-phosphofructo-2-kinase and its role for the regulation of glycolysis under hypoosmotic conditions were investigated. 6-Phosphofructo-2-kinase was found to be phosphorylated in vitro by protein kinase C at serine 652 and thereby inactivated. Protein phosphatase 2A reversed the phosphorylative inhibition of the enzyme. When yeast cells were shifted to hypotonic media, 6-phosphofructo-2-kinase was found to be phosphorylated and inactivated. Under in vivo conditions, two phosphate residues were incorporated into the enzyme. One of them is bound to serine 652, indicating that this modification was probably caused by yeast protein kinase C1. The second phosphate is bound to Ser8 within the N-terminal peptide T(1-41) which contains several serine residues but no protein kinase C recognition sequence. Site-directed mutagenesis confirmed that the phosphorylation of serine 652 but not the N-terminal modification is responsible for the in vivo inactivation of 6-phosphofructo-2-kinase. The obtained results suggest that the phosphorylation of 6-phosphofructo-2-kinase mediates a response of the cells to an activation of the hypoosmolarity MAP kinase pathway. Via a suppression of glycolysis, the inactivation of 6-phosphofructo-2-kinase is expected to be responsible for the observed accumulation of glucose 6-phosphate, an essential precursor of the cell wall glucans, and the decrease of glycerol, an important osmolyte.
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PMID:Phosphorylation and inactivation of yeast 6-phosphofructo-2-kinase contribute to the regulation of glycolysis under hypotonic stress. 1172 81

Rapamycin binds and inhibits the Tor protein kinases, which function in a nutrient-sensing signal transduction pathway that has been conserved from the yeast Saccharomyces cerevisiae to humans. In yeast cells, the Tor pathway has been implicated in regulating cellular responses to nutrients, including proliferation, translation, transcription, autophagy, and ribosome biogenesis. We report here that rapamycin inhibits pseudohyphal filamentous differentiation of S. cerevisiae in response to nitrogen limitation. Overexpression of Tap42, a protein phosphatase regulatory subunit, restored pseudohyphal growth in cells exposed to rapamycin. The tap42-11 mutation compromised pseudohyphal differentiation and rendered it resistant to rapamycin. Cells lacking the Tap42-regulated protein phosphatase Sit4 exhibited a pseudohyphal growth defect and were markedly hypersensitive to rapamycin. Mutations in other Tap42-regulated phosphatases had no effect on pseudohyphal differentiation. Our findings support a model in which pseudohyphal differentiation is controlled by a nutrient-sensing pathway involving the Tor protein kinases and the Tap42-Sit4 protein phosphatase. Activation of the MAP kinase or cAMP pathways, or mutation of the Sok2 repressor, restored filamentation in rapamycin treated cells, supporting models in which the Tor pathway acts in parallel with these known pathways. Filamentous differentiation of diverse fungi was also blocked by rapamycin, demonstrating that the Tor signaling cascade plays a conserved role in regulating filamentous differentiation in response to nutrients.
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PMID:The TOR signal transduction cascade controls cellular differentiation in response to nutrients. 1173 4

Protein phosphatase 2A (PP2A) is a multimeric serine/threonine phosphatase that carries out multiple functions. Although numerous observations suggest that PP2A plays a major role in downregulation of the mitogen-activated protein (MAP) kinase pathway, the precise mechanisms are unknown. To clarify the role of PP2A in growth factor (insulin, epidermal growth factor [EGF], and insulin-like growth factor 1 [IGF-1]) stimulation of the Ras/MAP kinase pathway, simian virus 40 small t antigen was expressed in Rat-1 fibroblasts which overexpress insulin receptors. Small t antigen is known to specifically inhibit PP2A by binding to the A PP2A regulatory subunit, interfering with the ability of PP2A to bind to its cellular substrates. Overexpressed small t protein was coimmunoprecipitated with PP2A and inhibited cellular PP2A activity but did not inhibit protein phosphatase 1 (PP1) activity. Insulin, IGF-1, and EGF stimulation also inhibited PP2A activity. Growth factor-stimulated Ras, Raf-1, MAP kinase, and mitogen-activated extracellular-signal-regulated kinase kinase (MEK) activities were elevated in small-t-antigen-expressing cells. Furthermore, Shc tyrosine phosphorylation and its association with Grb2 were also elevated in small-t-antigen-expressing cells. Expression levels of Shc, Ras, MEK, or MAP kinase and phosphorylation of insulin, EGF, and IGF-1 receptors were not altered. Interestingly, we found that PP2A associated with Shc in the basal state and dissociated in response to insulin and EGF and that this dissociation was inhibited by 65% in small-t-antigen-expressing cells. In addition, we found that PP2A associates with the phosphotyrosine-binding domain (PTB domain) of Shc and that phosphorylation of tyrosine 317 of Shc was required for PP2A-Shc dissociation. We conclude (i) that PP2A negatively regulates the Ras/MAP kinase pathway by binding to Shc, inhibiting tyrosine phosphorylation; (ii) that the Shc-PP2A association is mediated by the Shc PTB domain but the interaction is independent of phosphotyrosine binding, indicating a new molecular function for the PTB domain; (iii) that growth factor stimulation, or small-t-antigen expression, causes dissociation of the PP2A-Shc complex, facilitating Shc phosphorylation and downstream activations of the Ras/MAP kinase pathway; and (iv) that this defines a new mechanism of small-t-antigen action to promote mitogenesis.
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PMID:Protein phosphatase 2A forms a molecular complex with Shc and regulates Shc tyrosine phosphorylation and downstream mitogenic signaling. 1188 20

Individual subunits of protein phosphatase 2A (PP2A), protein phosphatase 4, and protein phosphatase 5 were knocked out in Drosophila Schneider 2 cells by using RNA interference. Ablation of either the scaffold (A) or catalytic (C) subunits of PP2A caused the disappearance of all PP2A subunits. Treating cells with double-stranded RNA targeting all four of the Drosophila PP2A regulatory subunits caused the disappearance of both the A and C subunits. The loss of PP2A subunits was associated with decreased protein stability indicating that only the heterotrimeric forms of PP2A are stable in intact cells. Ablation of total PP2A by using double-stranded RNA against either the A or C subunit, or specific ablation of the R2/B regulatory subunit, enhanced insulin-induced ERK activation. These results indicated that the R2/B subunit targets PP2A to the mitogen-activated protein (MAP) kinase cascade in Schneider 2 cells, where it acts as a negative regulator. A severe loss of viability occurred in cells in which total PP2A or both isoforms of the Drosophila R5/B56 subunit had been ablated. The reduced viability of these cells correlated with the induction of markers of apoptosis including membrane blebbing and stimulation of caspase-3-like activity. These observations indicated that PP2A has a powerful antiapoptotic activity that is specifically mediated by the R5/B56 regulatory subunits. In contrast to PP2A, ablation of protein phosphatase 4 caused only a slight reduction in cell growth but had no effect on MAP kinase signaling or apoptosis. Depletion of protein phosphatase 5 had no effects on MAP kinase, cell growth, or apoptosis.
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PMID:Actions of PP2A on the MAP kinase pathway and apoptosis are mediated by distinct regulatory subunits. 1190 83

Calcineurin (CN), a highly conserved Ca2+/calmodulin-regulated phosphatase, is a critical component of many calcium-regulated processes in mammalian cells, including T cell activation, cardiac hypertrophy, learning and memory. CN is specifically inhibited by the immunosuppressant drugs cyclosporin A and tacrolimus (FK506), and these drugs have served as valuable reagents in identifying the role of CN in a wide variety of cell types. CN may have additional functions in other cell types, and the loss of these functions may contribute to the side effects of these drugs, which include nephrotoxicity and neurotoxicity. A better understanding of the biological roles of CN in different cell types may promote the development of improved strategies for immunosuppression. We have been studying the CN signal transduction pathway in fission yeast because this system is amenable to genetics and has many advantages in terms of relevance to higher systems. Fission yeast has a single gene encoding the catalytic subunit of CN, ppb1+, that is essential for cytokinesis. We have shown that in fission yeast CN plays an essential role in maintaining chloride ion homeostasis and acts antagonistically with the Pmk1 MAP kinase pathway. We also carried out an isolation and a screening for several FK506-sensitive mutants in order to identify genes that share an essential function for viability with CN. Possible roles of these gene products in cellular functions in relation to calcineurin are discussed.
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PMID:[Functional analysis of calcineurin-mediated signalling pathway using fission yeast as a model system]. 1191 17


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