Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.7.11.11 (AMPK)
 12,425 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Saccharomyces cerevisiae neutral trehalase, encoded by NTH1, controls trehalose hydrolysis in response to multiple stress conditions, including nutrient limitation. The presence of three stress responsive elements (STREs, CCCCT) in the NTH1 promoter suggested that the transcriptional activator proteins Msn2 and Msn4, as well as the cAMP-dependent protein kinase (PKA), control the stress-induced expression of Nth1. Here, we give direct evidence that Msn2/Msn4 and the STREs control the heat-, osmotic stress- and diauxic shift-dependent induction of Nth1. Disruption of MSN2 and MSN4 abolishes or significantly reduces the heat- and NaCl-induced increases in Nth1 activity and transcription. Stress-induced increases in activity of a lacZ reporter gene put under control of the NTH1 promoter is nearly absent in the double mutant. In all instances, basal expression is also reduced by about 50%. The trehalose concentration in the msn2 msn4 double mutant increases less during heat stress and drops more slowly during recovery than in wild-type cells. This shows that Msn2/Msn4-controlled expression of enzymes of trehalose synthesis and hydrolysis help to maintain trehalose concentration during stress. However, the Msn2/Msn4-independent mechanism exists for heat control of trehalose metabolism. Site-directed mutagenesis of the three STREs (CCCCT changed to CATCT) in NTH1 promoter fused to a reporter gene indicates that the relative proximity of STREs to each other is important for the function of NTH1. Elimination of the three STREs abolishes the stress-induced responses and reduces basal expression by 30%. Contrary to most STRE-regulated genes, the PKA effect on the induction of NTH1 by heat and sodium chloride is variable. During diauxic growth, NTH1 promoter-controlled reporter activity strongly increases, as opposed to the previously observed decrease in Nth1 activity, suggesting a tight but opposite control of the enzyme at the transcriptional and post-translational levels. Apparently, inactive trehalase is accumulated concomitant with the accumulation of trehalose. These results might help to elucidate the general connection between control by STREs, Msn2/Msn4 and PKA and, in particular, how these components play a role in control of trehalose metabolism.
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PMID:Induction of neutral trehalase Nth1 by heat and osmotic stress is controlled by STRE elements and Msn2/Msn4 transcription factors: variations of PKA effect during stress and growth. 1065

Saccharomyces cerevisiae neutral trehalase (encoded by NTH1) is regulated by cAMP-dependent protein kinase (PKA) and by an endogenous modulator protein. A yeast strain with knockouts of CMK1 and CMK2 genes (cmk1cmk2) and its isogenic control (CMK1CMK2) were used to investigate the role of CaM kinase II in the in vitro activation of neutral trehalase during growth on glucose. In the exponential growth phase, cmk1cmk2 cells exhibited basal trehalase activity and an activation ratio by PKA very similar to that found in CMK1CMK2 cells. At diauxie, even though both cells presented comparable basal trehalase activities, cmk1cmk2 cells showed reduced activation by PKA and lower total trehalase activity when compared to CMK1CMK2 cells. To determine if CaM kinase II regulates NTH1 expression or is involved in post-translational modulation of neutral trehalase activity, NTH1 promoter activity was evaluated using an NTH1-lacZ reporter gene. Similar beta-galactosidase activities were found for CMK1CMK2 and cmk1cmk2 cells, ruling out the role of CaM kinase II in NTH1 expression. Thus, CaM kinase II should act in concert with PKA on the activation of the cryptic form of neutral trehalase. A model for trehalase regulation by CaM kinase II is proposed whereby the target protein for Ca2+/CaM-dependent kinase II phosphorylation is not the neutral trehalase itself. The possible identity of this target protein with the recently identified trehalase-associated protein YLR270Wp is discussed.
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PMID:Evidence for a modulation of neutral trehalase activity by Ca2+ and cAMP signaling pathways in Saccharomyces cerevisiae. 1174 9

Neutral trehalase from Neurospora crassa was expressed in Escherichia coli as a polypeptide of approximately 84 kDa in agreement with the theoretical size calculated from the corresponding cDNA. The recombinant neutral trehalase, purified by affinity chromatography exhibited a specific activity of 80-150 mU/mg protein. Optima of pH and temperature were 7.0 and 30 degrees C, respectively. The enzyme was absolutely specific for trehalose, and was quite sensitive to incubation at 40 degrees C. The recombinant enzyme was totally dependent on calcium, and was inhibited by ATP, copper, silver, aluminium and cobalt. K(M) was 42 mM, and V(max) was 30.6 nmol of glucose/min. The recombinant protein was phosphorylated by cAMP-dependent protein kinase, but not significantly activated. Immunoblotting with polyclonal antiserum prepared against the recombinant protein showed that neutral trehalase protein levels increased during exponential phase of N. crassa growth and dropped at the stationary phase. This is the first report of a neutral trehalase produced in E. coli with similar biochemical properties described for fungi native neutral trehalases, including calcium-dependence.
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PMID:Heterologous expression in Escherichia coli of Neurospora crassa neutral trehalase as an active enzyme. 1907 63

Trehalose is an alpha, alpha-1, 1-linked glucose disaccharide. In plants, trehalose is synthesized in two steps. Firstly, trehalose-6-phosphate synthase (TPS) converts UDP-glucose and glucose-6-phosphate to trehalose-6-phosphate (T6P); secondly, T6P-phosphatase (TPP) converts T6P into trehalose and Pi. Trehalose is further cleaved into glucose by trehalase. In extracts of most plants, including Arabidopsis, levels of both trehalose and T6P are low, nearing detection limits, and this has delayed research into their function. Trehalose is transported widely in plants, but transport of T6P is not thought to occur except possibly at the subcellular level. Feeding trehalose to Arabidopsis seedlings alters carbon allocation with massive starch accumulation in cotyledons and leaves and absence of starch and growth in shoot and root apices.The Arabidopsis genome has experienced extensive radiation of genes likely encoding enzymes of T6P metabolism: 4 and 10 genes are found with homology to TPS and TPP respectively and 7 genes are found with homology to both TPS and TPP. Complementation of Saccharomyces cerevisiae mutants has shown that AtTPS1, AtTPPA and AtTPPB are functional enzymes. In contrast just a single gene encoding a protein with trehalase activity has been found. Whilst most TPS proteins appear cytosolic, strikingly, some TPPs appear targeted to chloroplasts; trehalase on the other hand is extracellular. Transporters of trehalose and T6P have yet to be described. Arabidopsis tps1 mutants are embryo lethal and results suggest that T6P is essential for several other steps in development including root growth and floral transition. Accordingly, altering T6P content has a profound effect on plant habitus and impacts metabolite profiles, sugar utilization and photosynthesis. These large effects have hindered dissection of cause and effect. In contrast, plants with large alterations in sucrose-6-phosphate concentrations are indistinguishable from wild type, suggesting very different functions for these compounds. Recently, T6P at low micromolar concentrations has been shown in vitro and in vivo to inhibit SnRK1 of the SNF1/AMPK group of protein kinases. This supports a function for T6P as a sugar signaling molecule integrating metabolism and development in plants in relation to carbon supply.Genetic engineering of Arabidopsis as well as tobacco, potato and rice with TPS or TPS/TPP protein fusions reveals that trehalose metabolism also mediates multiple abiotic stress tolerances. Trehalose applications also mediate biotic stress resistances. Both Escherichia coli and Saccharomyces cerevisiae TPS/TPP protein fusions can be used to engineer stress tolerance suggesting that metabolites rather than proteins of the trehalose pathway are key stress tolerance elicitors. Results underscore the central role of trehalose metabolites in integrating carbon metabolism and stress responses with plant development.
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PMID:Trehalose Metabolites in Arabidopsis-elusive, active and central. 2230 48

Mutants with specific lesions were used to differentiate between the functions of glycogen and trehalose in S. cerevisiae. Diploids which harbor the glc1/glc1 mutation depend upon the phosphorylated, less active form of glycogen synthase and show a more active, phosphorylated form, of the enzyme trehalase. These conditions are due to a lesion in the regulating subunit of the cAMP-dependent protein kinase. Such cells are unable to sporulate. Diploids which contain the sst1/sst1 mutation have normal glycogen metabolism but their trehalose-6-phosphate synthase is not active. Such strains sporulate but germination is poor and only one-spore tetrads are formed. These results confirm that glycogen is needed to trigger sporulation while trehalose plays a role in the germination process. Different systems, I and II, of trehalose accumulation were proposed. System I would require the UDPG-linked trehalose synthase, whereas system II would constitute an alternative pathway, specifically induced or activated by the expression of a MAL gene. The presence of system II in its constitutive form in the constructed diploids would favour trehalose synthesis during growth on glucose, however, it did not overcome the glycogen deficiency during sporulation nor the lack of trehalose for germination. It seems that only system I, namely trehalose 6-P-synthase, plays a role in the germination process.
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PMID:Trehalose: Its role in germination of Saccharomyces cerevisiae. 2417 21


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