Chill-induced inhibition of photosynthesis: genotypic variation within Cucumis sativus

Cucumber is generally a thermophilic species; however, cultivars have been selected for higher yield during winter cultivation in unheated glasshouses in temperate regions. We tested whether photosynthesis in these varieties had greater chilling tolerance. There was no difference in the instantaneous reduction of photosynthesis at low temperature between four winter glasshouse and four summer field cultivars. After 5 d of 10 degrees C and 100 micro mol m(-2) s(-1) photon flux density, the four field cultivars had a sustained depression of photosynthesis after returning to clement conditions. This inhibition was associated with reduced rates of CO(2) fixation and photosystem II (PSII) electron transport in the light, but not with sustained PSII photoinhibition. However, photosynthesis in the glasshouse genotypes was nearly identical to the pre-chill rates. Chill impacts on light-adapted chlorophyll fluorescence parameters, such as the quantum yield of PSII electron transport (phi(PSII)), correlated well with overall photosynthesis. This demonstrates the potential for using these fast and non-invasive techniques to screen for chill-tolerant genotypes, with the potential to further improve winter cucumber yield in unheated glasshouses.     

Glyphosate-resistant goosegrass. Identification of a mutation in the target enzyme 5-enolpyruvylshikimate-3-phosphate synthase

The spontaneous occurrence of resistance to the herbicide glyphosate in weed species has been an extremely infrequent event, despite over 20 years of extensive use. Recently, a glyphosate-resistant biotype of goosegrass (Eleusine indica) was identified in Malaysia exhibiting an LD(50) value approximately 2- to 4-fold greater than the sensitive biotype collected from the same region. A comparison of the inhibition of 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) activity by glyphosate in extracts prepared from the resistant (R) and sensitive (S) biotypes revealed an approximately 5-fold higher IC(50)(glyphosate) for the (R) biotype. Sequence comparisons of the predicted EPSPS mature protein coding regions from both biotypes revealed four single-nucleotide differences, two of which result in amino acid changes. One of these changes, a proline to serine substitution at position 106 in the (R) biotype, corresponds to a substitution previously identified in a glyphosate-insensitive EPSPS enzyme from Salmonella typhimurium. Kinetic data generated for the recombinant enzymes suggests that the second substitution identified in the (R) EPSPS does not contribute significantly to its reduced glyphosate sensitivity. Escherichia coli aroA- (EPSPS deficient) strains expressing the mature EPSPS enzyme from the (R) biotype exhibited an approximately 3-fold increase in glyphosate tolerance relative to strains expressing the mature EPSPS from the (S) biotype. These results provide the first evidence for an altered EPSPS enzyme as an underlying component of evolved glyphosate resistance in any plant species.     

How to grab a microtubule on the move

A recent paper in Cell highlights a second mechanism of regulation of separase in addition to the bound inhibitor securin. This second pathway involves separase phosphorylation and is dependent on CDC2.     

Detecting single molecules launched from liquid surfaces in a time-of-flight mass spectrometer using ultraviolet and infrared lasers

The launch of molecules from liquid surfaces in a time of flight mass spectrometer has been investigated using different sample preparation techniques, and by exposing the liquid samples to two different laser wavelengths, 337 nm from a N2 ultraviolet laser and 10.6 microm from a CO2 infrared laser. The molecules were detected with cryodetectors measuring the energy of the individual molecules. We present insulin and lysozyme results from samples introduced into the vacuum through a micromachined silicon injector, and from samples consisting of a glycerol droplet deposited directly on the sample holder at the high voltage stage of the ion optics.     

Differences between rats and mice in induction of 4S beta-naphthoflavone-binding protein expression by treatment with beta-naphthoflavone

Sprague-Dawley rats and several inbred strains of mice were compared with respect to the induction of 4S BNF-binding protein expression by treatment with beta-naphthoflavone (BNF), an aryl hydrocarbon receptor (AhR) ligand. Inbred strains of mice chosen for the study encompassed 3 allelic forms of AhR found in Mus musculus so far. As it was reported by us earlier, treating rats with BNF caused a significant induction of BNF-binding protein expression. By contrast, no BNF-binding protein was detected in mice treated with BNF in corn oil as a vehicle or with corn oil itself.     

The 1.5 A crystal structure of a highly selected antiviral T cell receptor provides evidence for a structural basis of immunodominance

Despite a potential repertoire of >10(15) alphabeta T cell receptors (TcR), the HLA B8-restricted cytolytic T cell response to a latent antigen of Epstein-Barr virus (EBV) is strikingly limited in the TcR sequences that are selected. Even in unrelated individuals this response is dominated by a single highly restricted TcR clonotype that selects identical combinations of hypervariable Valpha, Vbeta, D, J, and N region genes. We have determined the 1.5 A crystal structure of this "public" TcR, revealing that five of the six hypervariable loops adopt novel conformations providing a unique combining site that contains a deep pocket predicted to overlay the HLA B8-peptide complex. The findings suggest a structural basis for the immunodominance of this clonotype in the immune response to EBV.     

EPR study of non-covalent spin labeled serum albumin and hemoglobin

Electron Paramagnetic Resonance (EPR) was used to investigate the Tempyo spin label (3-carbamoyl-2,2,5,5-tetramethyl-3-pyrrolin-1-yloxy) as a report group for the interactions and the conformational changes of lyophilized bovine serum albumin (BSA) and bovine hemoglobin (BH), as function of pH values in the range 2.5-11. The EPR spectra are similar with those of other non-covalently spin label porphyrins in frozen solution at very low temperatures. This behavior indicated a possible spin-spin interaction between the hemic iron and the nitroxide group. The changes in the EPR spectra as function of the pH are discussed in terms of conformational changes of the proteins. Spectral simulations and magnetic EPR parameters reveal the following: (i) one single paramagnetic species, with Gaussian line shape, was used for the best fits of experimental spectra in the case of serum albumin samples; and (ii) a weighted sum of Lorentzian and Gaussian line shape in the case of hemoglobin samples. The representation of correlation time vs. pH, reveals a dependence of degree of immobilization of spin label on the conformational changes of proteins in acidic and basic environment.     

Exercise,free radicals,and lipid peroxidation in type 1 diabetes mellitus

Indirect biochemical techniques have solely been used to ascertain whether type 1 diabetes mellitus patients are more susceptible to resting and exercise-induced oxidative stress. To date there is no direct evidence to support the contention that type 1 diabetic patients have increased levels of free radical species. Thus, the aim of this study was to use electron spin resonance (ESR) spectroscopy in conjunction with alpha-phenyl-tert-butylnitrone (PBN) spin trapping to measure pre- and postexercise free radical concentration in the venous blood of young male patients with type 1 diabetes mellitus (HbA(1c) = 8.2 +/- 1%, n = 12) and healthy matched controls (HbA(1c) = 5.5 +/- 0.2%, n = 13). Supporting measures of lipid peroxidation (malondialdehyde and lipid hydroperoxides), ambient blood glucose and selected antioxidants were also measured. The diabetic patients presented with a comparatively greater concentration of free radicals as measured by ESR and lipid hydroperoxides (LH) compared to the healthy group (p <.05, pooled rest and exercise data), although there was no difference in malondialdehyde (MDA) concentration. alpha-Tocopherol was comparatively lower in the healthy group (p <.05, pooled rest and exercise data vs. diabetic group) due to a selective decrease during physical exercise (p <.05 vs. rest). The hyperfine coupling constants recorded from the ESR spectra (a(Nitrogen) = 1.37 mT and abeta(Hydrogen) = 0.17 mT) are suggestive of either oxygen or carbon-centered species and are consistent with literature values. We suggest that the greater concentration of oxidants seen in the diabetic group may be due to increased glucose autoxidation as a function of this pathology and/or a lower exercise-induced oxidation rate of the major lipid soluble antioxidant alpha-tocopherol. We suggest that the ESR-detected radicals are secondary species derived from decomposition of LH because these are the major initial reaction products of free radical attack on cell membranes.