The Drosophila Melanogaster RAD54 Homolog, DmRAD54, Is Involved in the Repair of Radiation Damage and Recombination

The RAD54 gene of Saccharomyces cerevisiae plays a crucial role in recombinational repair of double-strand breaks in DNA. Here the isolation and functional characterization of the RAD54 homolog of the fruit fly Drosophila melanogaster, DmRAD54, are described. The putative Dmrad54 protein displays 46 to 57% identity to its homologs from yeast and mammals. DmRAD54 RNA was detected at all stages of fly development, but an increased level was observed in early embryos and ovarian tissue. To determine the function of DmRAD54, a null mutant was isolated by random mutagenesis. DmRAD54-deficient flies develop normally, but the females are sterile. Early development appears normal, but the eggs do not hatch, indicating an essential role for DmRAD54 in development. The larvae of mutant flies are highly sensitive to X rays and methyl methanesulfonate. Moreover, this mutant is defective in X- ray-induced mitotic recombination as measured by a somatic mutation and recombination test. These phenotypes are consistent with a defect in the repair of double-strand breaks and imply that the RAD54 gene is crucial in repair and recombination in a multicellular organism. The results also indicate that the recombinational repair pathway is functionally conserved in evolution

Effects of a defective ERAD pathway on growth and heterologous protein production in Aspergillus niger

Endoplasmic reticulum associated degradation (ERAD) is a conserved mechanism to remove misfolded proteins from the ER by targeting them to the proteasome for degradation. To assess the role of ERAD in filamentous fungi, we have examined the consequences of disrupting putative ERAD components in the filamentous fungus Aspergillus niger. Deletion of derA, doaA, hrdC, mifA, or mnsA in A. niger yields viable strains, and with the exception of doaA, no significant growth phenotype is observed when compared to the parental strain. The gene deletion mutants were also made in A. niger strains containing single- or multicopies of a glucoamylase–glucuronidase (GlaGus) gene fusion. The induction of the unfolded protein response (UPR) target genes (bipA and pdiA) was dependent on the copy number of the heterologous gene and the ERAD gene deleted. The highest induction of UPR target genes was observed in ERAD mutants containing multiple copies of the GlaGus gene. Western blot analysis revealed that deletion of the derA gene in the multicopy GlaGus overexpressing strain resulted in a 6-fold increase in the intracellular amount of GlaGus protein detected. Our results suggest that impairing some components of the ERAD pathway in combination with high expression levels of the heterologous protein results in higher intracellular protein levels, indicating a delay in protein degradation

Quantification of Grassland Biomass and Nitrogen Content through UAV Hyperspectral Imagery—Active Sample Selection for Model Transfer

Accurate retrieval of grassland traits is important to support management of pasture production and phenotyping studies. In general, conventional methods used to measure forage yield and quality rely on costly destructive sampling and laboratory analysis, which is often not viable in practical applications. Optical imaging systems carried as payload in Unmanned Aerial Vehicles (UAVs) platforms have increasingly been proposed as alternative non-destructive solutions for crop characterization and monitoring. The vegetation spectral response in the visible and near-infrared wavelengths provides information on many aspects of its composition and structure. Combining spectral measurements and multivariate modelling approaches it is possible to represent the often complex relationship between canopy reflectance and specific plant traits. However, empirical models are limited and strictly represent characteristics of the observations used during model training, therefore having low generalization potential. A method to mitigate this issue consists of adding informative samples from the target domain (i.e., new observations) to the training dataset. This approach searches for a compromise between representing the variability in new data and selecting only a minimal number of additional samples for calibration transfer. In this study, a method to actively choose new training samples based on their spectral diversity and prediction uncertainty was implemented and tested using a multi-annual dataset. Accurate predictions were obtained using hyperspectral imagery and linear multivariate models (Partial Least Squares Regression—PLSR) for grassland dry matter (DM; R2 = 0.92, RMSE = 3.25 dt ha−1), nitrogen (N) content in % of DM (R2 = 0.58, RMSE = 0.27%) and N-uptake (R2 = 0.91, RMSE = 6.50 kg ha−1). In addition, the number of samples from the target dates added to the training dataset could be reduced by up to 77% and 74% for DM and N-related traits, respectively, after model transfer. Despite this reduction, RMSE values for optimal transfer sets (identified after validation and used as benchmark) were only 20–30% lower than those values obtained after model transfer based on prediction uncertainty reduction, indicating that loss of accuracy was relatively small. These results demonstrate that considerably simple approaches based on UAV hyperspectral data can be applied in preliminary grassland monitoring frameworks, even with limited datasets

Haus Riswick grassland experiment with N fertilization and plant growth monitoring based on Unmanned Aerial Vehicle (UAV) hyperspectral imagery - campaigns of 2014 and 2017

Grassland (Lolium perenne) dry aboveground biomass, nitrogen content (N%) and N uptake (t ha-1) measured in experimental plots located near the city of Kleve, in Germany. These measurements correspond to five harvest dates, two in 2014 and three in 2017. In addition, UAV flights were performed on the same dates in the experimental site to acquire hyperspectral images, using the WageningenUR Hyperspectral Mapping System (HYMSY), a pushbroom imaging sensor comprising a spectrometer and a photogrammetric camera

A light-weight hyperspectral mapping system for unmanned aerial vehicles - The first results

Research opportunities using UAV remote sensing techniques are limited by the payload of the platform. Therefore small UAV's are typically not suitable for hyperspectral imaging due to the weight of the mapping system. In this research, we are developing a light-weight hyperspectral mapping system (< 2 kg) suitable to be mounted on small UAVs. The system is able to produce georeferenced and georectified hyperspectral data cubes in 400-1000nm spectral range at 10-50cm resolution. The georeferenced reflectance factor spectra cubes are to be used in e.g. precision agriculture and soil erosion research. In this paper we describe prototype of the system, the processing chain, and present preliminary results

Isolation of two laccase genes from the white-rot fungus Pleurotus eryngii and heterologous expression of the pel3 encoded protein

11 páginas, 4 figuras -- PAGS nros. 9-19In this paper we report the cloning and nucleotide sequence analysis of two new laccase genes from the white-rot fungus Pleurotus eryngii, named pel3 and pel4. Comparison of the protein sequences deduced from these genes with laccases previously described in P. eryngii indicates that these genes codify for new laccases in this fungus. We described the expression of pel3 gene in two different Aspergillus niger strains. Both the laccase signal peptide and the glucoamylase preprosequence of A. niger were used to target the secretion of the active enzyme. The highest levels of laccase expression were obtained by combining the last construction with an A. niger strain deficient in extracellular proteases secretion. The characterization of catalytic properties of the recombinant enzyme, together with the setting-up of a heterologous expression system for pel3, will provide the basis to study the biotechnological applications of this enzyme.Contract NMP2-CT-2006-026456 and the Spanish project BIO2003-00621 and S-0505/AMB0100. Financial support received from the “Comunidad de Madrid"Peer reviewe

Comparison Between the Performance of Quantitative Flow Ratio and Perfusion Imaging for Diagnosing Myocardial Ischemia

Objectives: This study compared the performance of the quantitative flow ratio (QFR) with single-photon emission computed tomography (SPECT) and positron emission tomography (PET) myocardial perfusion imaging (MPI) for the diagnosis of fractional flow reserve (FFR)−defined coronary artery disease (CAD). Background: QFR estimates FFR solely based on cine contrast images acquired during invasive coronary angiography (ICA). Head-to-head studies comparing QFR with noninvasive MPI are lacking. Methods: A total of 208 (624 vessels) patients underwent technetium- 99m tetrofosmin SPECT and [ 15O]H 2O PET imaging before ICA in conjunction with FFR measurements. ICA was obtained without using a dedicated QFR acquisition protocol, and QFR computation was attempted in all vessels interrogated by FFR (552 vessels). Results: QFR computation succeeded in 286 (52%) vessels. QFR correlated well with invasive FFR overall (R = 0.79; p < 0.001) and in the subset of vessels with an intermediate (30% to 90%) diameter stenosis (R = 0.76; p < 0.001). Overall, per-vessel analysis demonstrated QFR to exhibit a superior sensitivity (70%) in comparison with SPECT (29%; p < 0.001), whereas it was similar to PET (75%; p = 1.000). Specificity of QFR (93%) was higher than PET (79%; p < 0.001) and not different from SPECT (96%; p = 1.000). As such, the accuracy of QFR (88%) was superior to both SPECT (82%; p = 0.010) and PET (78%; p = 0.004). Lastly, the area under the receiver operating characteristics curve of QFR, in the overall sample (0.94) and among vessels with an intermediate lesion (0.90) was higher than SPECT (0.63 and 0.61; p < 0.001 for both) and PET (0.82; p < 0.001 and 0.77; p = 0.002), respectively. Conclusions: In this head-to-head comparative study, QFR exhibited a higher diagnostic value for detecting FFR-defined significant CAD compared with perfusion imaging by SPECT or PET

Phosphodiesterase 4 inhibition attenuates atrial natriuretic peptide-induced vascular hyperpermeability and loss of plasma volume

Inhibition of phosphodiesterase 4 (PDE4) to increase endothelial cAMP and stabilize the endothelial barrier attenuates acute inflammatory increases in vascular permeability. We extended this approach to attenuate physiological increases in vascular permeability in response to atrial natriuretic peptide (ANP), which acts with the kidney to regulate plasma volume. We measured blood-to-tissue albumin clearance and changes in plasma volume in isoflurane-anaesthetized mice (C57BL/6J) pre-treated with rolipram (8 mg kg−1i.p., 30 min). Rolipram significantly reduced albumin permeability, measured using a dual-label fluorescence method, in skin and skeletal muscle compared with ANP alone (500 ng kg−1 min−1). Skin and muscle tissue accounted for 70% of the reduction in whole body albumin clearance taking into account albumin clearance in gastrointestinal (GI) tissue, heart and kidney. The action of ANP and rolipram to modify albumin clearances in duodenum and jejunum could be accounted for by local increases in vascular perfusion to increase surface area for exchange. ANP increased haematocrit from 40.6% to 46.8%, corresponding to an average loss of 22% plasma fluid volume (227 μl), and this was almost completely reversed with rolipram. Renal water excretion accounted for less than 30% of plasma fluid loss indicating that reduced albumin permeability and reduced filtration into vasodilated GI tissue were the predominant actions of PDE4 inhibition. Similar fluid retention was measured in mice with endothelial-restricted deletion of the guanylyl cyclase-A receptor for ANP. Stabilizing the endothelial barrier to offset ANP-induced increases in vascular permeability may be part of a strategy to maintain plasma volume