A novel noninvasive procedure for high‐throughput screening of major seed traits

The large numbers of samples processed in breeding and biodiversity programmes require the development of efficient methods for the nondestructive evaluation of basic seed properties. Near‐infrared spectroscopy is the state‐of‐the‐art solution for this analytical demand, but it also has some limitations. Here, we present a novel, rapid, accurate procedure based on time domain‐nuclear magnetic resonance (TD‐NMR), designed to simultaneously quantify a number of basic seed traits without any seed destruction. Using a low‐field, benchtop ¹H‐NMR instrument, the procedure gives a high‐accuracy measurement of oil content (R² = 0.98), carbohydrate content (R² = 0.99), water content (R² = 0.98) and both fresh and dry weight of seeds/grains (R² = 0.99). The method requires a minimum of ~20 mg biomass per sample and thus enables to screen individual, intact seeds. When combined with an automated sample delivery system, a throughput of ~1400 samples per day is achievable. The procedure has been trialled as a proof of concept on cereal grains (collection of ~3000 accessions of Avena spp. curated at the IPK genebank). A mathematical multitrait selection approach has been designed to simplify the selection of outlying (most contrasting) accessions. To provide deeper insights into storage oil topology, some oat accessions were further analysed by three‐dimensional seed modelling and lipid imaging. We conclude that the novel TD‐NMR‐based screening tool opens perspectives for breeding and plant biology in general.     

Novel N-Acylethanolamide Derivatives Affect Body Weight and Energy Balance

Introduction – The obesity pandemic is multifactorial. Nutritional, pharmacologic and surgical interventions are limited in reach and efficacy, raising need for new therapeutics. Aims – Characterization of anorexigenic and cognitive effect and central mechanism of action of novel N-acylethanolamide derivatives. Methods – Sabra mice divided to similar experimental groups, injected IP with: oleyl-L-leucinolamide ( 1 A ), linoleyl-L-leucinolamide ( 4 A ), linoleyl-L-valinolamide ( 5 A ), oleyl-oxycarbonyl-L-valinolamide ( 1 B ), oleyl-oxycarbonyl-D-valinolamide ( 2 B ), oleylamine-carbonyl-L-valinolamide ( 3 B ), oleylamine-carbonyl-D-valinolamide ( 4 B ), and oleyl-L-hydroxyvalineamide ( 5 B ). Control group with vehicle. Body weight and food consumption followed for 39 days. Motor activity and cognitive function by open field test and eight-arm maze. Mice sacrificed and mechanism of action investigated by qPCR. The genes analyzed involved in energy balance and regulation of appetite. Catecholamines and serotonin evaluated. Results – Compounds 1 A , 5 A , 1 B – 4 B , caused significant weight loss of 4.2–5.6 % and 5 A , 1 B – 4 B , improved cognitive function following 8 i. p. injections of 1 mg/kg during 39 days, by different mechanisms. 5 A , 3 B and 4 B decreased food consumption, whereas 1 A , 5 A and 2 B increased motor activity. 1 A , 4 A , 1 B and 3 B elevated SIRT-1, associated with survival. POMC upregulated by 1 B and 2 B , CART by 1 B , 2 B and 1 A . NPY and CAMKK2 downregulated by 5 A . 4 B enhanced 5-HT levels. 4 A , 5 A , 1 B , 4 B , 5 B decreased FAAH, showing long lasting effect. Conclusions – These new compounds might be developed for the treatment of obesity and for improved cognitive function.     

Human base excision repair enzymes apurinic/apyrimidinic endonuclease1 (APE1), DNA polymerase beta and poly(ADP-ribose) polymerase 1: interplay between strand-displacement DNA synthesis and proofreading exonuclease activity

We examined interactions between base excision repair (BER) DNA intermediates and purified human BER enzymes, DNA polymerase β (pol β), apurinic/apyrimidinic endonuclease (APE1) and poly(ADP-ribose) polymerase-1 (PARP-1). Studies under steady-state conditions with purified BER enzymes and BER substrates have already demonstrated interplay between these BER enzymes that is sensitive to the respective concentrations of each enzyme. Therefore, in this study, using conditions of enzyme excess over substrate DNA, we further examine the question of interplay between BER enzymes on BER intermediates. The results reveal several important differences compared with data obtained using steady-state assays. Excess PARP-1 antagonizes the action of pol β, producing a complete block of long patch BER strand-displacement DNA synthesis. Surprisingly, an excess of APE1 stimulates strand-displacement DNA synthesis by pol β, but this effect is blocked by PARP-1. The APE1 exonuclease function appears to be modulated by the other BER proteins. Excess APE1 over pol β may allow APE1 to perform both exonuclease function and stimulation of strand-displacement DNA synthesis by pol β. This enables pol β to mediate long patch sub-pathway. These results indicate that differences in the stoichiometry of BER enzymes may regulate BER.     

Molecular chaperone RAP interacts with LRP1 in a dynamic bivalent mode and enhances folding of ligand-binding regions of other LDLR family receptors

The low-density lipoprotein receptor (LDLR) family of receptors are cell-surface receptors that internalize numerous ligands and play crucial role in various processes, such as lipoprotein metabolism, hemostasis, fetal development, etc. Previously, receptor-associated protein (RAP) was described as a molecular chaperone for LDLR-related protein 1 (LRP1), a prominent member of the LDLR family. We aimed to verify this role of RAP for LRP1 and two other LDLR family receptors, LDLR and vLDLR, and to investigate the mechanisms of respective interactions using a cell culture model system, purified system, and in silico modelling. Upon coexpression of RAP with clusters of the ligand-binding complement repeats (CRs) of the receptors in secreted form in insect cells culture, the isolated proteins had increased yield, enhanced folding, and improved binding properties compared with proteins expressed without RAP, as determined by circular dichroism and surface plasmon resonance. Within LRP1 CR-clusters II and IV, we identified multiple sites comprised of adjacent CR doublets, which provide alternative bivalent binding combinations with specific pairs of lysines on RAP. Mutational analysis of these lysines within each of isolated RAP D1/D2 and D3 domains having high affinity to LRP1 and of conserved tryptophans on selected CR-doublets of LRP1, as well as in silico docking of a model LRP1 CR-triplet with RAP, indicated a universal role for these residues in interaction of RAP and LRP1. Consequently, we propose a new model of RAP interaction with LDLR family receptors based on switching of the bivalent contacts between molecules over time in a dynamic mode.     

Alternative Splicing of Opioid Receptor Genes Shows a Conserved Pattern for 6TM Receptor Variants

Cellular and Molecular Neurobiology - The opioid receptor (OPR) family comprises the mu-, delta-, and kappa-opioid, and nociceptin receptors that belong to the superfamily of 7-transmembrane...     

A Preconscious Neural Mechanism of Hypnotically Altered Colors: A Double Case Study

Hypnotic suggestions may change the perceived color of objects. Given that chromatic stimulus information is processed rapidly and automatically by the visual system, how can hypnotic suggestions affect perceived colors in a seemingly immediate fashion? We studied the mechanisms of such color alterations by measuring electroencephalography in two highly suggestible participants as they perceived briefly presented visual shapes under posthypnotic color alternation suggestions such as “all the squares are blue”. One participant consistently reported seeing the suggested colors. Her reports correlated with enhanced evoked upper beta-band activity (22 Hz) 70–120 ms after stimulus in response to the shapes mentioned in the suggestion. This effect was not observed in a control condition where the participants merely tried to simulate the effects of the suggestion on behavior. The second participant neither reported color alterations nor showed the evoked beta activity, although her subjective experience and event-related potentials were changed by the suggestions. The results indicate a preconscious mechanism that first compares early visual input with a memory representation of the suggestion and consequently triggers the color alteration process in response to the objects specified by the suggestion. Conscious color experience is not purely the result of bottom-up processing but it can be modulated, at least in some individuals, by top-down factors such as hypnotic suggestions.     

The role of lipid‐related genes,aging‐related processes,and environment in healthspan

The inherent complexity of aging‐related traits can temper progress in unraveling the genetic origins of healthspan. We focus on two generations in the Framingham Heart Study, the original (FHS) and offspring (FHSO) cohorts, to determine whether aging‐related processes in changing environments can substantially impact the role of lipid‐related genes discovered in candidate gene (the apolipoprotein E (APOE) e2/3/4 polymorphism) and genome‐wide (the APOB rs1042034 (C/T)) studies, in regulation of total cholesterol (TC) and onset of cardiovascular disease (CVD). We demonstrate that the APOE e4 allele and APOB CC genotype can play detrimental, neutral, and protective sex‐specific roles in the etiology of CVD at different ages and in different environments. We document antagonistic roles for the e4 allele in the onset of CVD characterized by detrimental effects at younger ages (RR_{≤ 75 years} = 1.49, P = 7.5 × 10^?4) and protective effects at older ages (RR_76+years = 0.77, P = 0.044) for FHS participants. We found that disregarding the role of aging erroneously nullifies the significant effects of the e4 allele in this sample (RR = 0.92, P = 0.387). The leading biogenetic pathways mediating genetic effects on CVD may be more relevant to lipid metabolism for APOB than APOE. Aging‐related processes can modulate the strength of genetic associations with TC in the same individuals at different chronological ages. We found substantial differences in the effects of the same APOE and APOB alleles on CVD and TC across generations. The results suggest that aging‐related processes in changing environments may play key roles in the genetics of healthspan. Detailed systemic integrative analyses may substantially advance the progress.