Transport and metabolism of symplastic and apoplastic ascorbate during oxidative stress

Reactive oxygen species (ROS) are produced continuously in plants and act as important signalling molecules in many cellular processes including stress and defence responses. ROS can arise from external sources as well as being generated by the plant. Pollutants, such as ozone, enter the leaf via stomata and dissolve in the apoplast. ROS can arise both intracellularly and apoplastically: superoxide is produced during photosynthesis as well as by the plasma membrane bound NADPH oxidase during the oxidative burst. Under abiotic stresses such as drought or high light-intensity, superoxide production from photosynthetic electron flow is increased. Ascorbate plays a crucial role in symplastic and apoplastic ROS metabolism. Intracellular ascorbate metabolism is highly regulated; it is coupled to glutathione oxidation and reduction and is under tight enzymic control. Export of ascorbate into the apoplast increases during ozoneinduced stress. The apoplast redox state is considered to be more variable than the symplasm. Ascorbate is thought to be taken up from the apoplast in its oxidised form, DHA, via specific carriers, implying tight regulation of apoplastic/symplastic ascorbate transport. An apoplastic ascorbate breakdown pathway has recently been described by Green and Fry (2005). Ascorbate is oxidised and hydrolysed to yield oxalate via two novel intermediates, cyclic oxalyl L-threonate (cyc.ox.thr.) and 4-O-oxalyl-L-threonate (ox.thr.) A novel esterase is thought to catalyse the hydrolysis of ox.thr. to oxalate. Dehydro-L-ascorbate DHA was also hydrolysed to L-2,3-diketogulonate (DKG) which broke down to two unidentified compounds, C and E. It was not known whether this pathway operated intracellularly and how increased ROS production might affect flux through this pathway. The pathway, described, in the culture medium of 5-day-old rose cell suspension cultures but had not been investigated in planta. Intracellular and extracellular metabolism of [14C]ascorbate in [14C]ascorbate-loaded cells was investigated in response to oxidative stress induced by 0.1 and 1 mM H2O2 and 1 and 10 μM methyl viologen (MV2+). The symplasm became more oxidised in response to 0.1 mM H2O2; DHA levels increased and ascorbate decreased, but ox.thr. and oxalate, products of irreversible ascorbate breakdown, did not accumulate. Symplastic ox.thr. and oxalate accumulated in response to MV2+ and 1 mM H2O2. Ox.thr. and oxalate were observed in-planta. Flux through the pathway was increased in transgenic tobacco plants which overexpressed the cell wall-located enzyme ascorbate oxidase, suggesting that the redox state of the apoplast could increase apoplastic ascorbate breakdown via ox.thr. The rate of production of oxalate in vivo compared to in vitro studies suggested that the esterase was located to the symplasm as well as the apoplast. Oxalate did not appear to be metabolised further. Compounds C and E were neither observed in planta nor in 10-day old rose cell cultures. DKG and cyc.ox.thr. were present only in low levels. Export of 14C in [14C]ascorbate loaded cells increased in response to 1 and 5 mM H2O2. Increased export was characterised by a rapid response during the first 2 min of H2O2 exposure. In Arabidopsis and rose cell suspension cultures, export was often observed to occur in series of pulses. The amplitude of pulses increased within the first 2 min of H2O2 exposure. This was not thought to be a result of membrane disruption. 14C appeared to be exported as [14C]ascorbate and taken up as [14C]DHA, with minimal oxidation in the culture medium. These results provide more insight into intracellular ascorbate breakdown via ox.thr. and suggest that oxalate could accumulate in response to oxidative stress in plants. The export of ascorbate/DHA in pulses in response to H2O2 hints at novel mechanisms of regulation of ascorbate/DHA transport across the plasma membrane

A model for predicting dissolved organic carbon distribution in a reservoir water using fluorescence spectroscopy

A number of water treatment works (WTW) in the north of England (UK) have experienced problems in reducing the dissolved organic carbon (DOC) present in the water to a sufficiently low level. The problems are experienced in autumn/ winter when the colour increases and the coagulant dose at the WTW needs to be increased in order to achieve sufficient colour removal. However, the DOC content of the water varies little throughout the year. To investigate this further, the water was fractionated using resin adsorption techniques into its hydrophobic (fulvic and humic acid fractions) and hydrophilic (acid and non-acid fractions) components. The fractionation process yields useful information on the changing concentration of each fraction but is time consuming and labour intensive. Here, a method of rapidly determining fraction concentration was developed using fluorescence spectroscopy. The model created used synchronous spectra of fractionated material compared against bulk water spectra and predicted the fraction concentrations to within 10% for a specific water. The model was unable to predict fraction concentrations for waters from a different watershed

Spectral Libraries for SWATH-MS Assays for Drosophila melanogaster and Solanum lycopersicum.

Quantitative proteomics methods have emerged as powerful tools for measuring protein expression changes at the proteome level. Using MS-based approaches, it is now possible to routinely quantify thousands of proteins. However, prefractionation of the samples at the protein or peptide level is usually necessary to go deep into the proteome, increasing both MS analysis time and technical variability. Recently, a new MS acquisition method named SWATH is introduced with the potential to provide good coverage of the proteome as well as a good measurement precision without prior sample fractionation. In contrast to shotgun-based MS however, a library containing experimental acquired spectra is necessary for the bioinformatics analysis of SWATH data. In this study, spectral libraries for two widely used models are built to study crop ripening or animal embryogenesis, Solanum lycopersicum (tomato) and Drosophila melanogaster, respectively. The spectral libraries comprise fragments for 5197 and 6040 proteins for S. lycopersicum and D. melanogaster, respectively, and allow reproducible quantification for thousands of peptides per MS analysis. The spectral libraries and all MS data are available in the MassIVE repository with the dataset identifiers MSV000081074 and MSV000081075 and the PRIDE repository with the dataset identifiers PXD006493 and PXD006495

A new era of wide-field submillimetre imaging: on-sky performance of SCUBA-2

SCUBA-2 is the largest submillimetre wide-field bolometric camera ever built. This 43 square arc-minute field-of-view instrument operates at two wavelengths (850 and 450 microns) and has been installed on the James Clerk Maxwell Telescope on Mauna Kea, Hawaii. SCUBA-2 has been successfully commissioned and operational for general science since October 2011. This paper presents an overview of the on-sky performance of the instrument during and since commissioning in mid-2011. The on-sky noise characteristics and NEPs of the 450 and 850 micron arrays, with average yields of approximately 3400 bolometers at each wavelength, will be shown. The observing modes of the instrument and the on-sky calibration techniques are described. The culmination of these efforts has resulted in a scientifically powerful mapping camera with sensitivities that allow a square degree of sky to be mapped to 10 mJy/beam rms at 850 micron in 2 hours and 60 mJy/beam rms at 450 micron in 5 hours in the best weather.Comment: 18 pages, 15 figures.SPIE Conference series 8452, Millimetre, Submillimetre and Far-infrared Detectors and Instrumentation for Astronomy VI 201

The Effect of Galaxy Interactions on Molecular Gas Properties

© 2018. The American Astronomical Society. All rights reserved.Galaxy interactions are often accompanied by an enhanced star formation rate (SFR). Since molecular gas is essential for star formation, it is vital to establish whether and by how much galaxy interactions affect the molecular gas properties. We investigate the effect of interactions on global molecular gas properties by studying a sample of 58 galaxies in pairs and 154 control galaxies. Molecular gas properties are determined from observations with the JCMT, PMO, and CSO telescopes and supplemented with data from the xCOLD GASS and JINGLE surveys at 12CO(1-0) and 12CO(2-1). The SFR, gas mass (), and gas fraction (f gas) are all enhanced in galaxies in pairs by ∼2.5 times compared to the controls matched in redshift, mass, and effective radius, while the enhancement of star formation efficiency (SFE ≡SFR/) is less than a factor of 2. We also find that the enhancements in SFR, and f gas, increase with decreasing pair separation and are larger in systems with smaller stellar mass ratio. Conversely, the SFE is only enhanced in close pairs (separation <20 kpc) and equal-mass systems; therefore, most galaxies in pairs lie in the same parameter space on the SFR- plane as controls. This is the first time that the dependence of molecular gas properties on merger configurations is probed statistically with a relatively large sample and a carefully selected control sample for individual galaxies. We conclude that galaxy interactions do modify the molecular gas properties, although the strength of the effect is dependent on merger configuration.Peer reviewedFinal Accepted Versio

Ethylene Receptors, CTRs and EIN2 Target Protein Identification and Quantification Through Parallel Reaction Monitoring During Tomato Fruit Ripening.

Ethylene, the plant ripening hormone of climacteric fruit, is perceived by ethylene receptors which is the first step in the complex ethylene signal transduction pathway. Much progress has been made in elucidating the mechanism of this pathway, but there is still a lot to be done in the proteomic quantification of the main proteins involved, particularly during fruit ripening. This work focuses on the mass spectrometry based identification and quantification of the ethylene receptors (ETRs) and the downstream components of the pathway, CTR-like proteins (CTRs) and ETHYLENE INSENSITIVE 2 (EIN2). We used tomato as a model fruit to study changes in protein abundance involved in the ethylene signal transduction during fruit ripening. In order to detect and quantify these low abundant proteins located in the membrane of the endoplasmic reticulum, we developed a workflow comprising sample fractionation and MS analysis using parallel reaction monitoring. This work shows the feasibility of the identification and absolute quantification of all seven ethylene receptors, three out of four CTRs and EIN2 in four ripening stages of tomato. In parallel, gene expression was analyzed through real-time qPCR. Correlation between transcriptomic and proteomic profiles during ripening was only observed for three of the studied proteins, suggesting that the other signaling proteins are likely post-transcriptionally regulated. Based on our quantification results we were able to show that the protein levels of SlETR3 and SlETR4 increased during ripening, probably to control ethylene sensitivity. The other receptors and CTRs showed either stable levels that could sustain, or decreasing levels that could promote fruit ripening

12 co (3–2) high-resolution survey (cohrs) of the galactic plane: complete data release

We present the full data release of the 12CO (3–2) High-Resolution Survey (COHRS), which has mapped the inner Galactic plane over the range of 9.°5 ≤ l ≤ 62.°3 and ∣b∣ ≤ 0.°5. COHRS has been carried out using the Heterodyne Array Receiver Program on the 15 m James Clerk Maxwell Telescope in Hawaii. The released data are smoothed to have a spatial resolution of 16.″6 and a velocity resolution of 0.635 km s−1, achieving a mean rms of ∼0.6 K on TA* . The COHRS data are useful for investigating detailed three-dimensional structures of individual molecular clouds and large-scale structures such as spiral arms in the Galactic plane. Furthermore, data from other available public surveys of different CO isotopologues and transitions with similar angular resolutions to this survey, such as FUGIN, SEDIGISM, and CHIMPS/CHIMPS2, allow studies of the physical properties of molecular clouds and comparison of their states. In this paper, we report further observations on the second release and improved data reduction since the original COHRS release. We discuss the characteristics of the COHRS data and present integrated-emission images and a position–velocity (PV) map of the region covered. The PV map shows a good match with spiral-arm traces from existing CO and H i surveys. We also obtain and compare integrated one-dimensional distributions of 12CO (1–0) and (3–2) and those of star-forming populations