MAP kinase Hog1 mediates a cytochrome P450 oxidoreductase to promote the Sporisorium scitamineum cell survival under oxidative stress.

The MAP kinase high osmolarity glycerol 1 (Hog1) plays a central role in responding to external oxidative stress in budding yeast Saccchromyces cerevisiae. However, the downstream responsive elements regulated by Hog1 remain poorly understood. In this study, we report that a Sporisorium scitamineum orthologue of Hog1, named as SsHog1, induced transcriptional expression of a putative cytochrome P450 oxidoreductase encoding gene SsCPR1, to antagonize oxidative stress. We found that upon exposure to hydrogen peroxide (H2 O2 ), SsHog1 underwent strikingly phosphorylation, which was proved to be critical for transcriptional induction of SsCPR1. Loss of SsCPR1 led to hypersensitive to oxidative stress similar as the sshog1Δ mutant did, but was resistant to osmotic stress, which is different from the sshog1Δ mutant. On the other hand, overexpression of SsCPR1 in the sshog1Δ mutant could partially restore its ability of oxidative stress tolerance, which indicated that the Hog1 MAP kinase regulates the oxidative stress response specifically through cytochrome P450 (SsCpr1) pathway. Overall, our findings highlight a novel MAPK signalling pathway mediated by Hog1 in regulation of the oxidative stress response via the cytochrome P450 system, which plays an important role in host-fungus interaction.

Potassium Ferrate(VI) as a Highly Efficient and Environmentally Friendly Chemiluminescence Reagent in Acidic Solution.

Herein we report that the reactions of potassium ferrate (VI) with a number of reductants can produce strong chemiluminescence (CL) in acidic aqueous solution. The CL Spectra were registered and compared with the classical KMnO4 and NaClO-H2O2 CL systems. The characteristic emission peaks at 1268 and 1050 nm were observed, which are consistent to the spectrum obtained from the NaClO-H2O2 system. Additional emission bands at 680 nm further confirmed the formation of singlet oxygen dimers. The high CL intensity and the chemically green nature of K2FeO4, prompt us to further develop it as a novel CL reagent. Sensitive response and wide calibration ranges were obtained for dopamine, ascorbic acid, and ethanol. The linear range for the determination of three analytes were 50 nM to 50 µM for dopamine (LOD: 20 nM), 5.0 µM to 1.0 mM for ascorbic acid (LOD: 2.21 µM), and 0.5 µM to 1.0 mM for ethanol (LOD: 0.30 µM). Thus, K2FeO4 has a great potential for the postcolumn detection of those UV featureless compounds.

Arabidopsis AtPAP1 transcription factor induces anthocyanin production in transgenic Taraxacum brevicorniculatum.

KEY MESSAGE: This study developed a new purple coloured Taraxacum brevicorniculatum plant through genetic transformation using the Arabidopsis AtPAP1 gene, which overproduced anthocyanins in its vegetative tissues. Rubber-producing Taraxacum plants synthesise high-quality natural rubber (NR) in their roots and so are a promising alternative global source of this raw material. A major factor in its commercialization is the need for multipurpose exploitation of the whole plant. To add value to the aerial tissues, red/purple plants of the rubber-producing Taraxacum brevicorniculatum species were developed through heterologous expression of the production of anthocyanin pigment 1 (AtPAP1) transcription factor from Arabidopsis thaliana. The vegetative tissue of the transgenic plants showed an average of a 48-fold increase in total anthocyanin content over control levels, but with the exception of pigmentation, the transgenic plants were phenotypically comparable to controls and displayed similar growth vigor. Southern blot analysis confirmed that the AtPAP1 gene had been integrated into the genome of the high anthocyanin Taraxacum plants. The AtPAP1 expression levels were estimated by quantitative real-time PCR and were highly correlated with the levels of total anthocyanins in five independent transgenic lines. High levels of three cyanidin glycosides found in the purple plants were characterized by high performance liquid chromatography-mass spectrum analysis. The presence of NR was verified by NMR and infrared spectroscopy, and confirmed that NR biosynthesis had not been affected in the transgenic Taraxacum lines. In addition, other major phenylpropanoid products such as chlorogenic acid and quercetin glycosides were also enhanced in the transgenic Taraxacum. The red/purple transgenic Taraxacum lines described in this study would increase the future application of the species as a rubber-producing crop due to its additional health benefits.

Ring-Core Fluxgate Sensor for High Operation Temperatures up to 220 °C.

Fluxgate sensors are key devices for magnetic field surveys in geophysics. In areas such as deep drilling, fluxgate sensors may have to operate steadily at high temperatures for a prolonged period of time. We present an accordant ring-core type fluxgate sensor that is stable up to 220 °C. The high temperature consistency is achieved by using an Fe-based nanocrystalline magnetic core, PEEK structural components, an epoxy resin wrapping, as well as a broadband short-circuited working mode. The sensor was characterized at various temperatures up to 220 °C by evaluating impedance, hysteresis, permeability and sensitivity. We found a sensitivity of approximately 24 kV/T at 25 °C with an acceptable temperature coefficient of 742 ppm/°C throughout the range. The variation law of magnetic characteristics and their influence mechanism on output amplitude and phase are discussed.

Electrochemical Detection of Phosphate Ion in Body Fluids with a Magnesium Phosphate Modified Electrode.

An electrochemical sensor for phosphate detection in body fluids was developed based on the hydration transition of magnesium hydrogen phosphate (newberyite, MgHPO4·3H2O). The sensor was fabricated through incubation of a multi-walled carbon nanotube/Nafion (MWCNT/Nafion) modified glassy carbon electrode (GCE) in magnesium phosphate solution, where MgHPO4·3H2O was self-assembled on the electrode surface (denoted as MgP/MWCNT/Nafion). An electrooxidation peak at 1.0 V vs. Ag/AgCl was observed when the as-prepared electrode was subjected to a differential pulse voltammetry (DPV) scan in the presence of phosphate in acetate solution. When the DPV scan was performed in 0.4 - 1.3 V vs. Ag/AgCl, a linear relationship was observed between the peak height and the phosphate concentration in the range from 0.01 to 25 µM in the presence of 0.1 mM Mg2+ in the acetate solution with a limit of detection of 32 nM. And the sensor was successfully applied for phosphate detection in human urine and saliva samples with recoveries of 94.7 - 104.4 and 96 - 103.3%, respectively.

Histidine Kinase Sln1 and cAMP/PKA Signaling Pathways Antagonistically Regulate Sporisorium scitamineum Mating and Virulence via Transcription Factor Prf1.

Many prokaryotes and eukaryotes utilize two-component signaling pathways to counter environmental stress and regulate virulence genes associated with infection. In this study, we identified and characterized a conserved histidine kinase (SsSln1), which is the sensor of the two-component system of Sln1-Ypd1-Ssk1 in Sporisorium scitamineum. SsSln1 null mutant exhibited enhanced mating and virulence capabilities in S. scitamineum, which is opposite to what has been reported in Candida albicans. Further investigations revealed that the deletion of SsSLN1 enhanced SsHog1 phosphorylation and nuclear localization and thus promoted S. scitamineum mating. Interestingly, SsSln1 and cAMP/PKA signaling pathways antagonistically regulated the transcription of pheromone-responsive transcription factor SsPrf1, for regulating S. scitamineum mating and virulence. In short, the study depicts a novel mechanism in which the cross-talk between SsSln1 and cAMP/PKA pathways antagonistically regulates mating and virulence by balancing the transcription of the SsPRF1 gene in S. scitamineum.

Direct Electrochemical Sensing of Phosphate in Aqueous Solutions Based on Phase Transition of Calcium Phosphate.

Electrochemical determination of phosphate in aqueous solutions attracts considerable interests in both biological and environmental fields. Because of the electrochemically inactive nature of phosphate, direct electrochemical detection of phosphate is still a highly challenging task. Herein, we reported a direct electrochemical approach for the determination of phosphate based on the oxidation of coordinated OH during the phase transition of calcium phosphates (CaPs). The mixture of amorphous CaPs and octacalcium phosphate (Ca8(HPO4)2(PO4)4·5H2O), which acts as the starting material for hydroxyapatite (Ca10(PO4)6(OH)2), was self-assembled on a Nafion-modified glassy carbon electrode. The as-prepared electrode (CaPs/Nafion) showed a distinct oxidation peak at 1.0 V versus Ag/AgCl in phosphate solution. The peak heights were directly proportional to the concentration of phosphate from 0.1 to 10 µM in the presence of 1 mM Ca2+. After comprehensive characterization of the CaPs/Nafion electrode, it was understood that phosphate ions as a proton acceptor could stimulate the generation of coordinated OH from coordinated water (H2O) in CaP. The addition of Ca2+ could magnify the coordinated H2O source because of its hydration to H2O. The CaPs/Nafion electrode also displayed good selectivity as the electrochemical oxidization response was not affected by up to 10 µM of potentially competitive species like CO32-, NO3-, CH3COO-, SO42-, and Cl-. The results obtained in this work not only provided a new method for direct detection of phosphate in aqueous solution but also suggested that Ca2+ could be a promoter for electrochemical oxygen generation.

The Farnesyltransferase ß-Subunit Ram1 Regulates Sporisorium scitamineum Mating, Pathogenicity and Cell Wall Integrity.

The basidiomycetous fungus Sporisorium scitamineum causes a serious sugarcane smut disease in major sugarcane growing areas. Sexual mating is essential for infection to the host; however, its underlying molecular mechanism has not been fully studied. In this study, we identified a conserved farnesyltransferase (FTase) ß subunit Ram1 in S. scitamineum. The ram1Δ mutant displayed significantly reduced mating/filamentation, thus of weak pathogenicity to the host cane. The ram1Δ mutant sporidia showed more tolerant toward cell wall stressor Congo red compared to that of the wild-type. Transcriptional profiling showed that Congo red treatment resulted in notable up-regulation of the core genes involving in cell wall integrity pathway in ram1Δ sporidia compared with that of WT, indicating that Ram1 may be involved in cell wall integrity regulation. In yeast the heterodimeric FTase is responsible for post-translational modification of Ras (small G protein) and a-factor (pheromone). We also identified and characterized two conserved Ras proteins, Ras1 and Ras2, respectively, and a MAT-1 pheromone precursor Mfa1. The ras1Δ, ras2Δ and mfa1Δ mutants all displayed reduced mating/filamentation similar as the ram1Δ mutant. However, both ras1Δ and ras2Δ mutants were hypersensitive to Congo red while the mfa1Δ mutant was the same as wild-type. Overall our study displayed that RAM1 plays an essential role in S. scitamineum mating/filamentation, pathogenicity, and cell wall stability.

The MAP Kinase SsKpp2 Is Required for Mating/Filamentation in Sporisorium scitamineum.

In the phytopathogenic fungus Sporisorium scitamineum, sexual mating between two compatible haploid cells and the subsequent formation of dikaryotic hyphae is essential for infection. This process was shown to be commonly regulated by a mitogen-activated protein kinase (MAPK) and a cAMP/PKA signaling pathway in the corn smut fungus Ustilago maydis but remains largely unknown in S. scitamineum. In this study, we identified a conserved putative MAP kinase Kpp2 in S. scitamineum and named it as SsKpp2. The sskpp2Δ mutant displayed significant reduction in mating/filamentation, which could be partially restored by addition of cAMP or tryptophol, a quorum-sensing molecule identified in budding yeast. Transcriptional profiling showed that genes governing S. scitamineum mating or tryptophol biosynthesis were significantly differentially regulated in the sskpp2Δ mutant compared to the WT, under mating condition. Our results demonstrate that the MAP kinase SsKpp2 is required for S. scitamineum mating/filamentation likely through regulating the conserved pheromone signal transduction pathway and tryptophol production.

Nanomechanical Mapping of a Deformed Elastomer: Visualizing a Self-Reinforcement Mechanism.

Mapping the structure evolution and mechanical properties of elastic polymers or biomaterials during bulk deformation has been difficult, yet this information has long been thought to be key for understanding the structure-mechanical property relationship necessary to guide the design of new materials. Here we use a nanomechanical mapping to assess the structural evolution and mechanical properties of a deformed isoprene rubber (IR) to elucidate a self-reinforcement mechanism in this material. A hierarchical nanofibrillar structure, ranging from several to a hundred nanometers in size, comprised of fibers oriented parallel to the stretching direction was found. The nanofibers, connected by oriented amorphous tie chains, form a network structure that is responsible for significantly enhanced stress, a key factor giving rise to the self-reinforcement of IR and, more than likely, most elastomers that undergo strained-induced crystallization.