Intraspecfic variation in cold-temperature metabolic phenotypes of Arabidopsis lyrata ssp. petraea

Atmospheric temperature is a key factor in determining the distribution of a plant species. Alongside this, plant populations growing at the margin of their range may exhibit traits that indicate genetic differentiation and adaptation to their local abiotic environment. We investigated whether geographically separated marginal populations of Arabidopsis lyrata ssp. petraea have distinct metabolic phenotypes associated with exposure to cold temperatures. Seeds of A. petraea were obtained from populations along a latitudinal gradient, namely Wales, Sweden and Iceland and grown in a controlled cabinet environment. Mannose, glucose, fructose, sucrose and raffinose concentrations were different between cold treatments and populations, especially in the Welsh population, but polyhydric alcohol concentrations were not. The free amino acid compositions were population specific, with fold differences in most amino acids, especially in the Icelandic populations, with gross changes in amino acids, particularly those associated with glutamine metabolism. Metabolic fingerprints and profiles were obtained. Principal component analysis (PCA) of metabolite fingerprints revealed metabolic characteristic phenotypes for each population and temperature. It is suggested that amino acids and carbohydrates were responsible for discriminating populations within the PCA. Metabolite fingerprinting and profiling has proved to be sufficiently sensitive to identify metabolic differences between plant populations at different atmospheric temperatures. These findings show that there is significant natural variation in cold metabolism among populations of A. l. petraea which may signify plant adaptation to local climates. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Transport kinetics and selectivity of HpUreI, the urea channel from Helicobacter pylori

Helicobacter pylori's unique ability to colonize and survive in the acidic environment of the stomach is critically dependent on uptake of urea through the urea channel, HpUreI. Hence, HpUreI may represent a promising target for the development of specific drugs against this human pathogen. To obtain insight into the structure-function relationship of this channel, we developed conditions for the high-yield expression and purification of stable recombinant HpUreI. Detergent-solubilized HpUreI forms a homotrimer, as determined by chemical cross-linking. Urea dissociation kinetics of purified HpUreI were determined by means of the scintillation proximity assay, whereas urea efflux was measured in HpUreI-containing proteoliposomes using stopped-flow spectrometry to determine the kinetics and selectivity of the urea channel. The kinetic analyses revealed that urea conduction in HpUreI is pH-sensitive and saturable with a half-saturation concentration (or K(0.5)) of ~163 mM. The extent of binding of urea by HpUreI was increased at lower pH; however, the apparent affinity of urea binding (~150 mM) was not significantly pH-dependent. The solute selectivity analysis indicated that HpUreI is highly selective for urea and hydroxyurea. Removing either amino group of urea molecules diminishes their permeability through HpUreI. Similar to urea conduction, diffusion of water through HpUreI is pH-dependent with low water permeability at neutral pH.     

Growth stimulation and inhibition by salt in relation to Na<Superscript>+</Superscript> manipulating genes in xero-halophyte <Emphasis Type="Italic">Atriplex halimus</Emphasis> L.

In the present study, Na⁺ manipulating genes could contribute not only to ion homeostasis but also to growth stimulation with exposing the halophyte Atriplex halimus L. to moderate NaCl concentration. The stimulation of growth was attributed to Na⁺ accumulation inside the vacuole leading to increase leaf cell size as well as accelerate leaf cell division. Increasing the assimilatory surface could result in enhancing the photosynthetic rate. The reduction of A. halimus growth compared to optimum growth at 50 and 200 mM NaCl could be attributed to osmotic effect rather than the ionic one of salt stress. The inhibition of photosynthesis seemed to be resulted from limitation of CO₂ due to the osmotic effect on stomatal conductance rather than the activity loss of photosynthetic machinery. The depletion of starch content along with the increase in sucrose content could imply that photosynthesis may be a limiting for A. halimus growth. The fast coordinate induction of Na⁺ manipulating genes could reveal that the tolerance of A. halimus to high concentrations evolved from its ability to regulate and control Na⁺ influx and efflux. V-H ⁺-PPase may play a vital role in A. halimus tolerance to osmotic and/or ionic stress due to its kinetics of induction. It seemed that H⁺-ATPase plays a pivotal role in A. halimus tolerance to stress due to the increase in its protein level was detected with all NaCl concentrations as well as with PEG treatments. Both of these genes might be useful in improving stress tolerance in transgenic crops.     

From microbial gene essentiality to novel antimicrobial drug targets

Background

Bacterial respiratory tract infections, mainly caused by Streptococcus pneumoniae, Haemophilus influenzae and Moraxella catarrhalis are among the leading causes of global mortality and morbidity. Increased resistance of these pathogens to existing antibiotics necessitates the search for novel targets to develop potent antimicrobials.

Result

Here, we report a proof of concept study for the reliable identification of potential drug targets in these human respiratory pathogens by combining high-density transposon mutagenesis, high-throughput sequencing, and integrative genomics. Approximately 20% of all genes in these three species were essential for growth and viability, including 128 essential and conserved genes, part of 47 metabolic pathways. By comparing these essential genes to the human genome, and a database of genes from commensal human gut microbiota, we identified and excluded potential drug targets in respiratory tract pathogens that will have off-target effects in the host, or disrupt the natural host microbiota. We propose 249 potential drug targets, 67 of which are targets for 75 FDA-approved antimicrobials and 35 other researched small molecule inhibitors. Two out of four selected novel targets were experimentally validated, proofing the concept.

Conclusion

Here we have pioneered an attempt in systematically combining the power of high-density transposon mutagenesis, high-throughput sequencing, and integrative genomics to discover potential drug targets at genome-scale. By circumventing the time-consuming and expensive laboratory screens traditionally used to select potential drug targets, our approach provides an attractive alternative that could accelerate the much needed discovery of novel antimicrobials.

Electronic supplementary material

The online version of this article (doi:10.1186/1471-2164-15-958) contains supplementary material, which is available to authorized users.     

Improving Intra-Fractional Target Position Accuracy Using a 3D Surface Surrogate for Left Breast Irradiation Using the Respiratory-Gated Deep-Inspiration Breath-Hold Technique

Purpose

To evaluate the use of 3D optical surface imaging as a surrogate for respiratory gated deep-inspiration breath-hold (DIBH) for left breast irradiation.

Material and Methods

Patients with left-sided breast cancer treated with lumpectomy or mastectomy were selected as candidates for DIBH treatment for their external beam radiation therapy. Treatment plans were created on both free breathing (FB) and DIBH computed tomography (CT) simulation scans to determine dosimetric benefits from DIBH. The Real-time Position Management (RPM) system was used to acquire patient''s breathing trace during DIBH CT acquisition and treatment delivery. The reference 3D surface models from FB and DIBH CT scans were generated and transferred to the “AlignRT” system for patient positioning and real-time treatment monitoring. MV Cine images were acquired during treatment for each beam as quality assurance for intra-fractional position verification. The chest wall excursions measured on these images were used to define the actual target position during treatment, and to investigate the accuracy and reproducibility of RPM and AlignRT.

Results

Reduction in heart dose can be achieved using DIBH for left breast/chest wall radiation. RPM was shown to have inferior correlation with the actual target position, as determined by the MV Cine imaging. Therefore, RPM alone may not be an adequate surrogate in defining the breath-hold level. Alternatively, the AlignRT surface imaging demonstrated a superior correlation with the actual target positioning during DIBH. Both the vertical and magnitude real-time deltas (RTDs) reported by AlignRT can be used as the gating parameter, with a recommended threshold of ±3 mm and 5 mm, respectively.

Conclusion

The RPM system alone may not be sufficient for the required level of accuracy in left-sided breast/CW DIBH treatments. The 3D surface imaging can be used to ensure patient setup and monitor inter- and intra- fractional motions. Furthermore, the target position accuracy during DIBH treatment can be improved by AlignRT as a superior surrogate, in addition to the RPM system.     

Best of Both Worlds: Simultaneous High-Light and Shade-Tolerance Adaptations within Individual Leaves of the Living Stone Lithops aucampiae

“Living stones” (Lithops spp.) display some of the most extreme morphological and physiological adaptations in the plant kingdom to tolerate the xeric environments in which they grow. The physiological mechanisms that optimise the photosynthetic processes of Lithops spp. while minimising transpirational water loss in both above- and below-ground tissues remain unclear. Our experiments have shown unique simultaneous high-light and shade-tolerant adaptations within individual leaves of Lithops aucampiae. Leaf windows on the upper surfaces of the plant allow sunlight to penetrate to photosynthetic tissues within while sunlight-blocking flavonoid accumulation limits incoming solar radiation and aids screening of harmful UV radiation. Increased concentration of chlorophyll a and greater chlorophyll a∶b in above-ground regions of leaves enable maximum photosynthetic use of incoming light, while inverted conical epidermal cells, increased chlorophyll b, and reduced chlorophyll a∶b ensure maximum absorption and use of low light levels within the below-ground region of the leaf. High NPQ capacity affords physiological flexibility under variable natural light conditions. Our findings demonstrate unprecedented physiological flexibility in a xerophyte and further our understanding of plant responses and adaptations to extreme environments.