Identification of sodC encoding periplasmic [Cu,Zn]-superoxide dismutase in Salmonella

Abstract sodC , encoding [Cu,Zn]-cofactored Superoxide dismutase, once thought to be virtually confined to eukaryotes, has now been described in many Gram-negative pathogens that have their primary niche of colonization in the upper respiratory tract. Their role in host-parasite interactive biology is unknown. We here show that members of the major human and animal enteric pathogenic species Salmonella harbour a version of sodC most closely resembling that found in Brucella abortus . The enzyme it encodes is a novel candidate determinant of virulence in Salmonella , an intracellular pathogen potentially exposed to toxic oxygen free radicals within its intracellular niche.     

Activation of superoxide production and differential exocytosis in polymorphonuclear leukocytes by cytochalasins A,B, C,D and E: Effects of various ions

All of the common cytochalasins activate superoxide anion release and exocytosis of β-N-acetylglucosaminidase and lysozyme from guinea-pig polymorphonuclear leukocytes (neutrophils) incubated in a buffered sucrose medium. Half-maximal activation of both processes is produced by approx. 2 μM cytochalasin A, C >μM cytochalasin B ? 4–5 μM cytochalasin D, E. While maximal rates of O₂^? release and extents of exocytosis require extracellular calcium (1–2 mM), replacing sucrose with monovalent cation chlorides is inhibitory to neutrophil activation by cytochalasins. Na⁺, K⁺ or choline inhibited either cytochalasin B- or E-stimulated O₂^? production with IC₅₀ values of 5–10 mM and inhibition occurs whether Cl^?, NO₃^? or SCN^? is the anion added with Na⁺ or K⁺. Release of β-N-acetylglucosaminidase in control or cytochalasin B-stimulated cells is inhibited by NaCl (IC₅₀ ≈ 10 mM), while cytochalasin E-stimulated exocytosis is reduced less and K⁺ or choline chloride are ineffective in inhibiting either cytochalasin B- or E-stimulated exocytosis. Release of β-glucuronidase, myeloperoxidase or acid phosphatase from neutrophils incubated in buffered sucrose is not stimulated by cytochalasin B. Stimulation of either O₂^? or β-N-acetylglucosaminidase release by low concentrations of cytochalasin A is followed by inhibition of each at higher concentrations. It appears that all cytochalasins can activate both NAD(P)H oxidase and selective degranulation of neutrophils incubated in salt-restricted media and that differential inhibition of these two processes by monovalent cations and/or anions is produced at some step(s) subsequent to cytochalasin interaction with the cell.     

Vectorial redox reactions of physiological quinones. II. A study of transient semiquinone formation

Transient absorption changes during reduction of quinone in liposomes by external dithionite, in the absence and presence of initially trapped ferricyanide, were matched with absorption spectra of semiquinone and quinone in the blue region. Plastoquinone, ubiquinone-9 and phylloquinone, each having an isoprenoid side chain were compared with trimethyl-p-benzoquinone, ubiquinone-9 and menadione, which lack a long side chain.Semiquinone transients could only be observed by our spectroscopic technique during reduction of quinones lacking the chain. If Triton X-100 was added to the liposomes preparation semiquinone transients were also observed with the isoprenoid quinones. This result is consistent with the view that isoprenoid quinones build domains in the membranes, in which the life time of the semiquinone might be decreased by fast disproportionation, and to which dithionite has limited access.     

Temperature dependence of superoxide dismutase activity in plankton

The effect of temperature (from 1 to 37 °C) on in vitro effective superoxide dismutase (SOD) activity of several organisms was investigated and compared. Antarctic plankton, cultures of the alga Nannochloropsis sp., and the cyanobacterium Synechococcus strain WH 7803, and pure bovine erythrocyte SOD was studied. It was found that in all cases SOD activity increased with decreasing temperature within the temperature range assayed, in the Polar as well as the temperate plankton cells. This behavior of SOD is counterintuitive in terms of our experience when looking at enzyme activity or any other chemical reaction. We suggest a theoretical explanation for this apparently odd behavior. The advantage of such behavior is that the same amount of antioxidant will act better under low temperatures when reactive oxygen species (ROS) increase. Moreover, this protective process would act in vivo at a faster pace than the ex novo enzyme synthesis.     

Two modes of bicarbonate utilization in the marine green macroalga Ulva lactuca

The green marine macroalga Ulva lactuca L. was found to be able to utilize HCO₃^? from sea water in two ways. When grown in flowing natural sea water at 16°C under constant dim irradiance, photosynthesis at pH8.4 was suppressed by acetazolamide but unaffected by 4,4′-diisothiocyanostilbene-2,2′-disulphonate. These responses indicate that photosynthetic HCO₃^? utilization was via extracellular carbonic anhydrase (CA) -mediated dehydration followed by CO₂ uptake. The algae were therefore described as being in a ‘CA state’. If treated for more than 10 h in a sea water flow-through system at pH9.8, these thalli became insensitive to acetazolamide but sensitive to 4,4′-diisothiocyanostilbene-2,2′-disulphonate. This suggests the involvement of an anion exchanger (AE) in the direct uptake of HCO₃^?, and these plants were accordingly described as being in an ‘AE state’. Such thalli showed an approximately 10-fold higher apparent affinity for HCO₃^? (at pH9.4) than those in the ‘CA state’, while thalli of both states showed a very high apparent affinity for CO₂. These results suggest that the two modes of HCO₃^? utilization constitute two ways in which inorganic carbon may enter the Ulva lactuca cells, with the direct entry of HCO₃^?, characterizing the ‘AE state’, being inducible and possibly functioning as a complementary uptake system at high external pH values (e.g. under conditions conducive to high photosynthetic rates). Both mechanisms of entry appear to be connected to concentrating CO₂ inside the cell, probably via a separate mechanism operating intracellularly.     

DIA-Based Proteomics Identifies IDH2 as a Targetable Regulator of Acquired Drug Resistance in Chronic Myeloid Leukemia

Drug resistance is a critical obstacle to effective treatment in patients with chronic myeloid leukemia. To understand the underlying resistance mechanisms in response to imatinib mesylate (IMA) and adriamycin (ADR), the parental K562 cells were treated with low doses of IMA or ADR for 2 months to generate derivative cells with mild, intermediate, and severe resistance to the drugs as defined by their increasing resistance index. PulseDIA-based (DIA [data-independent acquisition]) quantitative proteomics was then employed to reveal the proteome changes in these resistant cells. In total, 7082 proteins from 98,232 peptides were identified and quantified from the dataset using four DIA software tools including OpenSWATH, Spectronaut, DIA-NN, and EncyclopeDIA. Sirtuin signaling pathway was found to be significantly enriched in both ADR-resistant and IMA-resistant K562 cells. In particular, isocitrate dehydrogenase (NADP(+)) 2 was identified as a potential drug target correlated with the drug resistance phenotype, and its inhibition by the antagonist AGI-6780 reversed the acquired resistance in K562 cells to either ADR or IMA. Together, our study has implicated isocitrate dehydrogenase (NADP(+)) 2 as a potential target that can be therapeutically leveraged to alleviate the drug resistance in K562 cells when treated with IMA and ADR.     

Efficacy of salicylic acid in modulating physiological andbiochemical mechanisms to improve postharvest longevity in cut spikes of Consolida ajacis (L.) Schur.

Postharvest losses of cut flowers is one of the considerable challenges restricting their efficient marketability. Consequently, such challenges have triggered a constant hunt for developing compatible postharvest treatments to mitigate postharvest losses. Interestingly, recent studies entrench extensive role of salicylic acid (SA) in mitigating postharvest losses in various flower systems. The current investigation focusses on role of SA in augmenting physiological and biochemical responses to mitigate postharvest senescence in cut spikes of Consolida ajacis. The cut spikes of C. ajacis were supplemented with various SA treatments viz, 2 mM, 4 mM, 6 mM. The effects of these treatments were evaluated against control set of spikes placed in distilled water. Our study indicates considerable increment in postharvest longevity of cut spikes, besides an increase in solution uptake, sugar and protein content of tepal tissues.SA augmented antioxidant system via upsurge in phenolic content and antioxidant enzymes viz, superoxide dismutase (SOD), catalase (CAT) and ascorbate peroxidase (APX) to forfend reactive oxygen species (ROS) related oxidative damage. SA profoundly reduced lipoxygenase (LOX) activity to preserve the membrane integrity and thus prevented seepage of solutes from tepal tissues. These results authenticate SA particularly 4 mM concentration as effective postharvest treatment to preserve the postharvest quality of C. ajacis cut spikes.     

Comparative study of the antioxidant activity of the essential oils of five plants against the H2O2 induced stress in Saccharomyces cerevisiae

The purpose of this work was to investigate the protective effect of five essential oils (EOs); Rosmarinus officinalis, Thymus vulgaris, Origanum compactum Benth., Eucalyptus globulus Labill. and Ocimum basilicum L.; against oxidative stress induced by hydrogen peroxide in Saccharomyces cerevisiae. The chemical composition of the EOs was analyzed by gas chromatography (GC) and gas chromatography-mass spectrometry (GC/MS). The in vitro antioxidant activity was evaluated and the protective effect of EOs was investigated. Yeast cells were pretreated with different concentrations of EOs (6.25–25 µg/ml) for an hour then incubated with H₂O₂ (2 mM) for an additional hour. Cell viability, antioxidants (Catalase, Superoxide dismutase and Glutathione reductase) and metabolic (Succinate dehydrogenase) enzymes, as well as the level of lipid peroxidation (LPO) and protein carbonyl content (PCO) were evaluated. The chemical composition of EOs has shown the difference qualitatively and quantitatively. Indeed, O. compactum mainly contained Carvacrol, O. basilicum was mainly composed of Linalool, T. vulgaris was rich in thymol, R. officinalis had high α-Pinene amount and for E. globulus, eucalyptol was the major compound. The EOs of basil, oregano and thyme were found to possess the highest amount of total phenolic compounds. Moreover, they have shown the best protective effect on yeast cells against oxidative stress induced by H₂O₂. In addition, in a dose dependent manner of EOs in yeast medium, treated cells had lower levels of LPO, lower antioxidant and metabolic enzymes activity than cells exposed to H₂O₂ only. The cell viability was also improved. It seems that the studied EOs are efficient natural antioxidants, which can be exploited to protect against damages and serious diseases related to oxidative stress.     

Inactivation of Rabbit Muscle Creatine Kinase by Hydrogen Peroxide

The effects of xanthine + xanthine oxidase-generated reactive oxygen species (ROS) on rabbit muscle creatine kinase (CK) were studied. Xanthine (0.1 mM) + xanthine oxidase (30 mU/ml) inhibited activity of rabbit muscle CK (1.2mU/ml). Catalase (100/ml), but not SOD (100 U/ml), deferoxamine (100μM) or mannitol (20 mM), protected CK from inactivation; suggesting that H₂O₂ was responsible for inactivation. These results were different from previously reported findings on bovine heart CK that superoxide radicals inactivate the enzyme. Thus, enzymes with homologous structures may have different reactivities to different ROS. H₂O₂-induced inactivation of rabbit muscle CK was accompanied by a decrease in its thiol group content, whereas no significant changes in the protein structure were detected by SDS-PAGE or carbonyl content. These results suggest that oxidation of -SH groups by H₂O₂ seems to be a major mechanism of activation of rabbit muscle CK by xanthine + xanthine oxidase. Such inactivation of CK by H₂O₂ may be important in ROS-induced pathology.     

The Growth,physiological and biochemical response of foxtail millet to atrazine herbicide

Foxtail millet (Pennisetum glaucum L.) is a vital crop that is planted as food and fodder crop around the globe. There is only limited information is present for abiotic stresses on the physiological responses to atrazine. A field experiment was conducted to investigate the effects of different atrazine dosages on the growth, fluorescence and physiological parameters i.e., malonaldehyde (MDA) and reactive oxygen species (ROS) (H₂O₂ and O₂) in the leaves to know the extent of atrazine on oxidative damage of foxtail millet. Our experiment consisted of 0, 2.5, 12.5, 22.5 and 32.5 (mg/kg) of labeled atrazine doses on 2 foxtaill millet varieties. High doses of atrazine significantly enhanced ROS and MDA synthesis in the plant leaves. Enzymes activities like ascorbate peroxidase (APX) and peroxidase (POD) activities enhanced, while catalase (CAD) and superoxide dismutase (SOD) activities reduced with increasing atrazine concentrations. Finally atrazine doses at 32.5 mg/kg reduced chlorophyll contents, while chlorophyll (a/b) ratio also enhanced. Biomass, plant height, chlorophyll fluorescence parameters, minimal and maximal fluorescence (Fo, Fm), maximum and actual quantum yield, photochemical quenching coefficient, and electron transport rate are decreased with increasing atrazine doses.