Feedback control of adrenal steroidogenesis via H2O2-dependent, reversible inactivation of peroxiredoxin III in mitochondria

Certain members of the peroxiredoxin (Prx) family undergo inactivation through hyperoxidation of the catalytic cysteine to sulfinic acid during catalysis and are reactivated by sulfiredoxin; however, the physiological significance of this reversible regulatory process is unclear. We now show that PrxIII in mouse adrenal cortex is inactivated by H(2)O(2) produced by cytochrome P450 enzymes during corticosterone production stimulated by adrenocorticotropic hormone. Inactivation of PrxIII triggers a sequence of events including accumulation of H(2)O(2), activation of p38 mitogen-activated protein kinase, suppression of steroidogenic acute regulatory protein synthesis, and inhibition of steroidogenesis. Interestingly, levels of inactivated PrxIII, activated p38, and sulfiredoxin display circadian oscillations. Steroidogenic tissue-specific ablation of sulfiredoxin in mice resulted in the persistent accumulation of inactive PrxIII and suppression of the adrenal circadian rhythm of corticosterone production. The coupling of CYP11B1 activity to PrxIII inactivation provides a feedback regulatory mechanism for steroidogenesis that functions independently of the hypothalamic-pituitary-adrenal axis.     

Identification of an ISR-related metabolite produced by Pseudomonas chlororaphis O6 against the wildfire pathogen pseudomonas syringae pv.tabaci in tobacco

Pseudomonas chlororaphis O6 exhibits induced systemic resistance (ISR) against P. syringae pv. tabaci in tobacco. To identify one of the ISR metabolites, O6 cultures were extracted with organic solvents, and the organic extracts were subjected to column chromatography followed by spectroscopy analyses. The ISR bioassay-guided fractionation was carried out for isolation of the metabolite. Highresolution mass spectrometric analysis of the metabolite found C(9)H(9)O(3)N with an exact mass of 179.0582. LC/MS analysis in positive mode showed an (M+H)(+) peak at m/zeta 180. Nuclear magnetic resonance ((1)H, (13)C) analyses identified all protons and carbons of the metabolite. Based on the spectroscopy data, the metabolite was identified 4-(aminocarbonyl) phenylacetate (4-ACPA). 4-ACPA applied at 68.0 mM exhibited ISR activity at a level similar 1.0 mM salicylic acid. This is the first report to identify an ISR metabolite produced by P. chlororaphis O6 against the wildfire pathogen P. syringae pv. tabaci in tobacco.     

Modulation of nitrosative stress by S-nitrosoglutathione reductase is critical for thermotolerance and plant growth in Arabidopsis

Nitric oxide (NO) is a key signaling molecule in plants. This analysis of Arabidopsis thaliana HOT5 (sensitive to hot temperatures), which is required for thermotolerance, uncovers a role of NO in thermotolerance and plant development. HOT5 encodes S-nitrosoglutathione reductase (GSNOR), which metabolizes the NO adduct S-nitrosoglutathione. Two hot5 missense alleles and two T-DNA insertion, protein null alleles were characterized. The missense alleles cannot acclimate to heat as dark-grown seedlings but grow normally and can heat-acclimate in the light. The null alleles cannot heat-acclimate as light-grown plants and have other phenotypes, including failure to grow on nutrient plates, increased reproductive shoots, and reduced fertility. The fertility defect of hot5 is due to both reduced stamen elongation and male and female fertilization defects. The hot5 null alleles show increased nitrate and nitroso species levels, and the heat sensitivity of both missense and null alleles is associated with increased NO species. Heat sensitivity is enhanced in wild-type and mutant plants by NO donors, and the heat sensitivity of hot5 mutants can be rescued by an NO scavenger. An NO-overproducing mutant is also defective in thermotolerance. Together, our results expand the importance of GSNOR-regulated NO homeostasis to abiotic stress and plant development.     

Transduced human PEP-1-heat shock protein 27 efficiently protects against brain ischemic insult

Reactive oxygen species contribute to the development of various human diseases. Ischemia is characterized by both significant oxidative stress and characteristic changes in the antioxidant defense mechanism. Heat shock protein 27 (HSP27) has a potent ability to increase cell survival in response to oxidative stress. In the present study, we have investigated the protective effects of PEP-1-HSP27 against cell death and ischemic insults. When PEP-1-HSP27 fusion protein was added to the culture medium of astrocyte and primary neuronal cells, it rapidly entered the cells and protected them against cell death induced by oxidative stress. Immunohistochemical analysis revealed that, when PEP-1-HSP27 fusion protein was intraperitoneally injected into gerbils, it prevented neuronal cell death in the CA1 region of the hippocampus in response to transient forebrain ischemia. Our results demonstrate that transduced PEP-1-HSP27 protects against cell death in vitro and in vivo, and suggest that transduction of PEP-1-HSP27 fusion protein provides a potential strategy for therapeutic delivery in various human diseases in which reactive oxygen species are implicated, including stroke.     

A new approach for on-line enrichment in electrophoresis of dilute protein solutions

A method is described for on-line enrichment/zone sharpening of a sample of negatively charged proteins (an analogous method for cationic proteins can be designed). The sample is applied on the top of a 5-mm thick layer of a neutral polyacrylamide gel which rests on another 5-mm thick, large-pore polyacrylamide gel which contains positively charged groups. The latter gel layer is attached to the neutral gel column, used for the electrophoretic separation of the proteins. When a voltage is applied the proteins start migrating and become electrostatically adsorbed at the top of the charged, large-pore gel layer (pH 5.4). With the upper electrode vessel filled with a buffer of a pH higher (pH 7.7) than that employed in the enrichment step and with a voltage between the electrodes, these enriched proteins are released (because the enrichment gel is non-charged at pH 7.7) with zone sharpening and migrate into the 5-cm long column (i.d. 5 mm) of a neutral, large-pore polyacrylamide gel for electrophoretic analysis. Upon the electrophoretic migration from the enrichment gel into the separation gel a second zone sharpening may occur, if the increase in pH from 5.4 to 7.7 in the separation gel is not close to momentary. By employing colored test proteins the efficiency of the enrichment step is visually illustrated by a picture. The principle of the concentration method described has been employed also in chromatographic experiments and can with appropriate modifications also be used in other electrophoretic methods, such as capillary electrophoresis.     

Social Facilitation of Synchronized Molting Behavior in the Spider Amaurobius ferox (Araneae, Amaurobiidae)

During the social period, molting behavior of the young spider, Amaurobius ferox, is highly synchronized within the clutch. Result of the experimental study suggests that social facilitation among group members increased the synchronization. The duration of the molting period of grouped spiderlings was significantly shorter than that of individually isolated spiderlings. Involving the particular maternal strategy in food supply, this phenomenon might have adaptive values in the maintenance of mutual tolerance among the siblings by decreasing the interindividual difference in development and in the avoidance of cannibalism on molting individuals. This probably will also serve to make the peaceful collective behaviors of the spiderlings in matriphagy and cooperative prey capture during their social period.     

Oral administration of a tri-therapy for central pattern generator activation in paraplegic mice: Proof-of-concept of efficacy

Spinal cord injury (SCI) is a neurological condition, for which no cure exists, typically leading to an immediate and irreversible loss of sensory and voluntary motor functions accompanied by significant health problems. We conducted proof-of-concept experiments aimed at assessing efficacy upon oral administration of a novel combination therapy for central pattern generator (CPG) activation and corresponding locomotor movement generation in completely paraplegic animals. Co-administration orally (by gavage) of buspirone, levodopa and carbidopa was found to dose-dependently induce episodes of steady weight-bearing stepping in low-thoracic (Th9/10) spinal cord-transected (Tx) mice (with no other form of assistance or training). Robust hindlimb stepping with weight-bearing capabilities was induced with the tri-therapy but not with clinically relevant doses of these compounds administered separately. These results provide evidence suggesting that this drug combination may be ideally suited to constitute a first-in-class therapy (CPG activator) for locomotor activity induction in chronic SCI individuals, given that efficacy was shown using commercially available brain-permeable small molecules, already known as safe for the treatment of various neurological indications.     

Theoretical calculation of the sugar concentrations during enzymatic production of fructooligosaccharides

Summary In a batch production of fructooligosaccharides from sucrose, the concentrations of residual sucrose, glucose and fructooligosaccharides at a given reaction time(t) and initial sucrose concentration(S₀) were theoretically calculated by the following correlation equations: Glucose(t) = 0.0653 S₀ × ln(t); Fructooligosaccharides(t) = 0.1636 S₀ × ln(t); Sucrose(t)=S₀ - Glucose(t) + FOS(t).