The effect of drought stress on heterozygosity–fitness correlations in pedunculate oak (Quercus robur)

Background and Aims

The interaction between forest fragmentation and predicted climate change may pose a serious threat to tree populations. In small and spatially isolated forest fragments, increased homozygosity may directly affect individual tree fitness through the expression of deleterious alleles. Climate change-induced drought stress may exacerbate these detrimental genetic consequences of forest fragmentation, as the fitness response to low levels of individual heterozygosity is generally thought to be stronger under environmental stress than under optimal conditions.

Methods

To test this hypothesis, a greenhouse experiment was performed in which various transpiration and growth traits of 6-month-old seedlings of Quercus robur differing in multilocus heterozygosity (MLH) were recorded for 3 months under a well-watered and a drought stress treatment. Heterozygosity–fitness correlations (HFC) were examined by correlating the recorded traits of individual seedlings to their MLH and by studying their response to drought stress.

Key Results

Weak, but significant, effects of MLH on several fitness traits were obtained, which were stronger for transpiration variables than for the recorded growth traits. High atmospheric stress (measured as vapour pressure deficit) influenced the strength of the HFCs of the transpiration variables, whereas only a limited effect of the irrigation treatment on the HFCs was observed.

Conclusions

Under ongoing climate change, increased atmospheric stress in the future may strengthen the negative fitness responses of trees to low MLH. This indicates the necessity to maximize individual multilocus heterozygosity in forest tree breeding programmes.     

Conformation-specific anti-Mad2 monoclonal antibodies for the dissection of checkpoint signaling

The spindle assembly checkpoint (SAC) ensures accurate chromosome segregation during mitosis by delaying the activation of the anaphase-promoting complex/cyclosome (APC/C) in response to unattached kinetochores. The Mad2 protein is essential for a functional checkpoint because it binds directly to Cdc20, the mitotic co-activator of the APC/C, thereby inhibiting progression into anaphase. Mad2 exists in at least 2 different conformations, open-Mad2 (O-Mad2) and closed-Mad2 (C-Mad2), with the latter representing the active form that is able to bind Cdc20. Our ability to dissect Mad2 biology in vivo is limited by the absence of monoclonal antibodies (mAbs) useful for recognizing the different conformations of Mad2. Here, we describe and extensively characterize mAbs specific for either O-Mad2 or C-Mad2, as well as a pan-Mad2 antibody, and use these to investigate the different Mad2 complexes present in mitotic cells. Our antibodies validate current Mad2 models but also suggest that O-Mad2 can associate with checkpoint complexes, most likely through dimerization with C-Mad2. Furthermore, we investigate the makeup of checkpoint complexes bound to the APC/C, which indicate the presence of both Cdc20-BubR1-Bub3 and Mad2-Cdc20-BubR1-Bub3 complexes, with Cdc20 being ubiquitinated in both. Thus, our defined mAbs provide insight into checkpoint signaling and provide useful tools for future research on Mad2 function and regulation.     

Influence of consumption of a high-protein vs. high-carbohydrate meal on the physiological cortisol and psychological mood response in men and women

Consumption of meals with different macronutrient contents, especially high in carbohydrates, may influence the stress-induced physiological and psychological response. The objective of this study was to investigate effects of consumption of a high-protein vs. high-carbohydrate meal on the physiological cortisol response and psychological mood response. Subjects (n = 38, 19 m/19f, age =25 ± 9 yrs, BMI = 25.0 ± 3.3 kg/m2) came to the university four times, fasted, for either condition: rest-protein, stress-protein, rest-carbohydrate, stress-carbohydrate (randomized cross-over design). Stress was induced by means of a psychological computer-test. The test-meal was either a high-protein meal (En% P/C/F 65/5/30) or a high-carbohydrate meal (En% P/C/F 6/64/30), both meals were matched for energy density (4 kJ/g) and daily energy requirements (30%). Per test-session salivary cortisol levels, appetite profile, mood state and level of anxiety were measured. High hunger, low satiety (81 ± 16, 12 ± 15 mm VAS) confirmed the fasted state. The stress condition was confirmed by increased feelings of depression, tension, anger, anxiety (AUC stress vs. rest p < 0.02). Consumption of the high-protein vs. high-carbohydrate meal did not affect feelings of depression, tension, anger, anxiety. Cortisol levels did not differ between the four test-sessions in men and women (AUC nmol·min/L p > 0.1). Consumption of the test-meals increased cortisol levels in men in all conditions (p < 0.01), and in women in the rest-protein and stress-protein condition (p < 0.03). Men showed higher cortisol levels than women (AUC nmol·min/L p < 0.0001). Consumption of meals with different macronutrient contents, i.e. high-protein vs. high-carbohydrate, does not influence the physiological and psychological response differentially. Men show a higher meal-induced salivary cortisol response compared with women.     

Recent Progress in Development of Tnt1 Functional Genomics Platform for Medicago truncatula and Lotus japonicus in Bulgaria

Legumes, as protein-rich crops, are widely used for human food, animal feed and vegetable oil production. Over the past decade, two legume species, Medicago truncatula and Lotus japonicus, have been adopted as model legumes for genomics and physiological studies. The tobacco transposable element, Tnt1, is a powerful tool for insertional mutagenesis and gene inactivation in plants. A large collection of Tnt1-tagged lines of M. truncatula cv. Jemalong was generated during the course of the project 'GLIP': Grain Legumes Integrated Project, funded by the European Union (www.eugrainlegumes.org). In the project 'IFCOSMO': Integrated Functional and COmparative genomics Studies on the MOdel Legumes Medicago truncatula and Lotus japonicus, supported by a grant from the Ministry of Education, Youth and Science, Bulgaria, these lines are used for development of functional genomics platform of legumes in Bulgaria. This review presents recent advances in the evaluation of the M. truncatula Tnt1 mutant collection and outlines the steps that are taken in using the Tnt1-tagging for generation of a mutant collection of the second model legume L. japonicus. Both collections will provide a number of legume-specific mutants and serve as a resource for functional and comparative genomics research on legumes. Genomics technologies are expected to advance genetics and breeding of important legume crops (pea, faba bean, alfalfa and clover) in Bulgaria and worldwide.     

Strigolactones fine-tune the root system

Strigolactones were originally discovered to be involved in parasitic weed germination, in mycorrhizal association and in the control of shoot architecture. Despite their clear role in rhizosphere signaling, comparatively less attention has been given to the belowground function of strigolactones on plant development. However, research has revealed that strigolactones play a key role in the regulation of the root system including adventitious roots, primary root length, lateral roots, root hairs and nodulation. Here, we review the recent progress regarding strigolactone regulation of the root system and the antagonism and interplay with other hormones.     

Naturally occurring Phe151Leu substitution near a conserved folding module lowers stability of glutathione transferase P1-1

Glutathione transferases (GSTs) are a family of enzymes that detoxify electrophilic compounds, such as carcinogens or drugs, by conjugating them to glutathione. The enzymes have contributed to the understanding of protein structure, due to large differences in amino acid sequence within the family, yet similar architecture and folding. Our objective was to conduct a systematic survey of GSTP1 polymorphisms and their function. Nearly all variants detected were known polymorphisms: IVS4+13C>A; Ile105Val; Ala114Val; and g.2596T>C (Ser185Ser). However, we also found a novel Phe151Leu substitution in an African-American subject (1 out of 111). Kinetic parameters for the conjugation reaction with 1-chloro-2,4-dinitrobenzene (CDNB) were determined for the novel variant enzyme purified via heterologous expression in Escherichia coli. Five substrates were used for measurement of specific activities, including isothiocyanate compounds that occur in cruciferous vegetables (benzylisothiocyanate, phenethylisothiocyanate, and sulforaphane). Such isothiocyanate substrates are potential cancer chemopreventive agents that are conjugated by GSTs. No major change in kinetic parameters was observed. However, the half-life at 50 degrees C of the Leu 151 enzyme was reduced to 12 min, as compared to 28 min for the Phe 151 enzyme. Residue 151 is located at the N-terminus of helix alpha6 in GST motif II, surrounded by hydrophobic residues, and near the conserved "hydrophobic staple" and N-capping box motifs. These local structural elements aid in formation of helix alpha6 and promote proper folding and protein stability. Analysis of the three-dimensional structure showed that substitution of Phe 151 with Leu produces a hydrophobic cavity in the GSTP1 core, thereby destabilizing its structure. Phe151Leu represents one of the first-described allelic variations in a protein folding motif.     

Individual variation in pup vocalizations and absence of behavioral signs of maternal vocal recognition in Weddell seals (Leptonychotes weddellii)

Individually stereotyped vocalizations often play an important role in relocation of offspring in gregarious breeders. In phocids, mothers often alternate between foraging at sea and attending their pup. Pup calls are individually distinctive in various phocid species. However, experimental evidence for maternal recognition is rare. In this study, we recorded Weddell seal (Leptonychotes weddellii) pup vocalizations at two whelping patches in Atka Bay, Antarctica, and explored individual vocal variation based on eight vocal parameters. Overall, 58% of calls were correctly classified according to individual. For males (n= 12) and females (n= 9), respectively, nine and seven individuals were correctly identified based on vocal parameters. To investigate whether mothers respond differently to calls of familiar vs. unfamiliar pups, we conducted playback experiments with 21 mothers. Maternal responses did not differ between playbacks of own, familiar, and unfamiliar pup calls. We suggest that Weddell seal pup calls may need to contain only a critical amount of individually distinct information because mothers and pups use a combination of sensory modalities for identification. However, it cannot be excluded that pup developmental factors and differing environmental factors between colonies affect pup acoustic behavior and the role of acoustic cues in the relocation process.     

Phthalates Are Metabolised by Primary Thyroid Cell Cultures but Have Limited Influence on Selected Thyroid Cell Functions In Vitro

Phthalates are plasticisers added to a wide variety of products, resulting in measurable exposure of humans. They are suspected to disrupt the thyroid axis as epidemiological studies suggest an influence on the peripheral thyroid hormone concentration. The mechanism is still unknown as only few in vitro studies within this area exist. The aim of the present study was to investigate the influence of three phthalate diesters (di-ethyl phthalate, di-n-butyl phthalate (DnBP), di-(2-ethylhexyl) phthalate (DEHP)) and two monoesters (mono-n-butyl phthalate and mono-(2-ethylhexyl) phthalate (MEHP)) on the differentiated function of primary human thyroid cell cultures. Also, the kinetics of phthalate metabolism were investigated. DEHP and its monoester, MEHP, both had an inhibitory influence on 3''-5''-cyclic adenosine monophosphate secretion from the cells, and MEHP also on thyroglobulin (Tg) secretion from the cells. Results of the lactate dehydrogenase-measurements indicated that the MEHP-mediated influence was caused by cell death. No influence on gene expression of thyroid specific genes (Tg, thyroid peroxidase, sodium iodine symporter and thyroid stimulating hormone receptor) by any of the investigated diesters could be demonstrated. All phthalate diesters were metabolised to the respective monoester, however with a fall in efficiency for high concentrations of the larger diesters DnBP and DEHP. In conclusion, human thyroid cells were able to metabolise phthalates but this phthalate-exposure did not appear to substantially influence selected functions of these cells.     

Genome sequence of the clover-nodulating Rhizobium leguminosarum bv. trifolii strain TA1

Rhizobium leguminosarum bv. trifolii strain TA1 is an aerobic, motile, Gram-negative, non-spore-forming rod that is an effective nitrogen fixing microsymbiont on the perennial clovers originating from Europe and the Mediterranean basin. TA1 however is ineffective with many annual and perennial clovers originating from Africa and America. Here we describe the features of R. leguminosarum bv. trifolii strain TA1, together with genome sequence information and annotation. The 8,618,824 bp high-quality-draft genome is arranged in a 6 scaffold of 32 contigs, contains 8,493 protein-coding genes and 83 RNA-only encoding genes, and is one of 20 rhizobial genomes sequenced as part of the DOE Joint Genome Institute 2010 Community Sequencing Program.