Tritium forms discrimination in ryegrass under constant tritium exposure: From seed germination to seedling autotrophy.

Uncertainties remain regarding the fate of atmospheric tritium after it has been assimilated in grasslands (ryegrass) in the form of TFWT (Tissue Free Water Tritium) or OBT (Organically Bound Tritium). One such uncertainty relates to the tritium forms discrimination during transfer from TFWT to OBT resulting from photosynthesis (OBTphoto), corresponding to the OBTphoto/TFWT ratio. In this study, the OBT/TFWT ratio is determined by experiments in the laboratory using a ryegrass model and hydroponic cultures, with constant activity of tritium in the form of tritiated water (denoted as HTO) in the "water" compartment (liquid HTO) and "air" compartment (HTO vapour in the air). The OBTphoto/TFWT ratio and the exchangeable OBT fraction are measured for three parts of the plant: the leaf, seed and root. Plant growth is modelled using dehydrated biomass measurements taken over time in the laboratory and integrating physiological functions of the plant during the first ten days after germination. The results suggest that there is no measurable discrimination of tritium in the plant organic matter produced by photosynthesis.

Fragile sites of 45S rDNA of Lolium multiflorum are not hotspots for chromosomal breakages induced by X-ray.

Sites of 45S rDNA of Lolium are regions denominated fragile sites (FSs), constituting regions slightly stained with DAPI due to increased DNA unpacking in metaphasic chromosomes. Considered to be fragile regions in the genome, the FSs might be more responsive to induced breaks and result in chromosomal fragments and rearrangements, unless repairing mechanisms such as recombination or de novo telomere formation play a role at the break site of the DNA. Thus, this study aimed at investigating if SFs from Lolium are hotspots for the occurrence of breakages induced by X-ray and if they are regions favorable to synthesize new telomeres, using Hordeum vulgare as a comparative model. Lolium multiflorum and H. vulgare seedlings were irradiated with 20 and 50 Gy X-ray and evaluated one day following the irradiation and at 7-days intervals for a 28-days period, using FISH technique with 45S rDNA and Arabidopsis-type telomere probes in order to investigate the presence of chromosomal breakages and new telomere formation. H. vulgare did not survive after a few days of irradiation due to the increased rate of abnormalities. L. multiflorum also exhibited chromosomal abnormalities following the exposure, yet over the 28-days trial it had a decrease in the chromosomal damage rate and formation of de novo telomere has not been detected along this time. Despite being considered to be fragile regions in the genome, the 45S rDNA sites of Lolium are not hotspots to chromosomal breakages after the induction of breakages.

DNA repair after X-irradiation: lessons from plants.

The effects of low-dose radiation causing DNA damage continue to be subjects of interest. Problems with existing approaches to low-dose DNA damage are that single-strand breaks (the predominant radiation-induced lesion) are very rapidly repaired and that results using current methods for measuring DNA damage can be difficult to interpret. As a novel approach, we conducted studies using plants (rye grass and the model plant Arabidopsis) exposed to X-rays and used the alkaline comet assay to measure DNA damage and repair after exposures. Consistent with previous studies, we detected so-called 'rapid' and 'slow' phases of DNA repair after acute exposures of 5 and 15 Gy. After exposures corresponding to 2 Gy and lower, 'rapid' repair was so fast that it was difficult to detect. We also found that the so-called 'slow' phase in both plants actually consisted of two components; an initial period of negligible repair lasting 80-120 min followed by a period of rapid repair lasting <30 min. Using Arabidopsis mutants homozygous for both ATM and BRCA1, we found that both of these genes are required for DNA repair during the 3-h period of our experiments, indicating that the 'slow' phase involves a homologous repair (HR) of double-strand breaks and clustered single-strand breaks. The lag of repair in the 'slow' phase presumably involves induction of expression of genes involved in HR repair such as BRCA1 and RAD51.

Latitudinal variation in ambient UV-B radiation is an important determinant of Lolium perenne forage production, quality, and digestibility.

Few studies to date have considered the responses of agriculturally important forage grasses to UV-B radiation. Yet grasses such as Lolium perenne have a wide current distribution, representing exposure to a significant variation in ambient UV-B. The current study investigated the responses of L. perenne (cv. AberDart) to a simulated latitudinal gradient of UV-B exposure, representing biologically effective UV-B doses at simulated 70, 60, 50, 40, and 30° N latitudes. Aspects of growth, soluble compounds, and digestibility were assessed, and results are discussed in relation to UV-B effects on forage properties and the implications for livestock and bio-ethanol production. Aboveground biomass production was reduced by approximately 12.67% with every 1 kJ m(-2) day(-1) increase in biologically weighted UV-B. As a result, plants grown in the highest UV-B treatment had a total biomass of just 13.7% of controls. Total flavonoids were increased by approximately 76% by all UV-B treatments, while hydroxycinnamic acids increased in proportion to the UV-B dose. Conversely, the digestibility of the aboveground biomass and concentrations of soluble fructans were reduced by UV-B exposure, although soluble sucrose, glucose, and fructose concentrations were unaffected. These results highlight the capacity for UV-B to directly affect forage productivity and chemistry, with negative consequences for digestibility and bioethanol production. Results emphasize the need for future development and distribution of L. perenne varieties to take UV-B irradiance into consideration.

UV responses of Lolium perenne raised along a latitudinal gradient across Europe: a filtration study.

Lolium perenne (cv. AberDart) was grown at 14 locations along a latitudinal gradient across Europe (37-68°N) to study the impact of ultraviolet radiation (UV) and climate on aboveground growth and foliar UV-B absorbing compounds. At each location, plants were grown outdoors for 5 weeks in a replicated UV-B filtration experiment consisting of open, UV-B transparent (cellulose diacetate) and UV-B opaque (polyester) environments. Fourier transform-infrared spectroscopy was used to compare plant metabolite profiles in relation to treatment and location. UV radiation and climatic parameters were determined for each location from online sources and the data were assessed using a combination of anova and multiple regression analyses. Most of the variation in growth between the locations was attributable to the combination of climatic parameters, with minimum temperature identified as an important growth constraint. However, no single environmental parameter could consistently account for the variability in plant growth. Concentrations of foliar UV-B absorbing compounds showed a positive trend with solar UV across the latitudinal gradient; however, this relationship was not consistent in all treatments. The most striking experimental outcome from this study was the effect of presence or absence of filtration frames on UV-absorbing compounds. Overall, the study demonstrates the value of an European approach in studying the impacts of natural UV across a large latitudinal gradient. We have shown the feasibility of coordinated UV filtration at multiple sites but have also highlighted the need for open controls and careful interpretation of plant responses.

Fluxes in central carbohydrate metabolism of source leaves in a fructan-storing C3 grass: rapid turnover and futile cycling of sucrose in continuous light under contrasted nitrogen nutrition status.

This work assessed the central carbohydrate metabolism of actively photosynthesizing leaf blades of a C3 grass (Lolium perenne L.). The study used dynamic (13)C labelling of plants growing in continuous light with contrasting supplies of nitrogen ('low N' and 'high N') and mathematical analysis of the tracer data with a four-pool compartmental model to estimate rates of: (i) sucrose synthesis from current assimilation; (ii) sucrose export/use; (iii) sucrose hydrolysis (to glucose and fructose) and resynthesis; and (iv) fructan synthesis and sucrose resynthesis from fructan metabolism. The contents of sucrose, fructan, glucose, and fructose were almost constant in both treatments. Labelling demonstrated that all carbohydrate pools were turned over. This indicated a system in metabolic steady state with equal rates of synthesis and degradation/consumption of the individual pools. Fructan content was enhanced by nitrogen deficiency (55 and 26% of dry mass at low and high N, respectively). Sucrose content was lower in nitrogen-deficient leaves (2.7 versus 6.7%). Glucose and fructose contents were always low (<1.5%). Interconversions between sucrose, glucose, and fructose were rapid (with half-lives of individual pools ranging between 0.3 and 0.8 h). Futile cycling of sucrose through sucrose hydrolysis (67 and 56% of sucrose at low and high N, respectively) and fructan metabolism (19 and 20%, respectively) was substantial but seemed to have no detrimental effect on the relative growth rate and carbon-use efficiency of these plants. The main effect of nitrogen deficiency on carbohydrate metabolism was to increase the half-life of the fructan pool from 27 to 62 h and to effectively double its size.

Dark-mediated dormancy release in stratified Lolium rigidum seeds is associated with higher activities of cell wall-modifying enzymes and an apparent increase in gibberellin sensitivity.

Dormancy release in freshly matured, imbibed annual ryegrass (Lolium rigidum) seeds is inhibited by light and involves a decrease in seed sensitivity to abscisic acid. Other processes involved in dormancy release in the dark were investigated by measuring seed storage compound mobilisation and the activity of cell wall-degrading enzymes. Activities of endo-ß-mannanase and total peroxidase were higher in dark-stratified compared to light-stratified seeds, indicating that weakening of the structures constraining the embryo was accelerated in the dark. A dramatic degradation of storage proteins in light-stratified seeds, accompanied by induction of a high molecular mass protease, suggests that maintenance of storage(-like) proteins is also important in dark-mediated dormancy release. α-Amylase activity was induced in dark-stratified seeds at least 48 h prior to radicle emergence upon transfer to conditions permitting germination, or in light-stratified seeds supplied with exogenous gibberellin A(4). This suggests that (a) α-amylase is involved in stimulation of germination of non-dormant L. rigidum seeds, and (b) dark-stratified seeds have an increased sensitivity to gibberellins which permits the rapid induction of α-amylase activity upon exposure to germination conditions. Overall, it appears that a number of processes, although possibly minor in themselves, occur in concert during dark-stratification to contribute to dormancy release.

Light and nitrogen competition limit Lolium perenne in experimental grasslands of increasing plant diversity.

Positive species richness effects on aboveground community productivity in experimental grasslands have been reported to correlate with variable responses of individual species. So far, it is largely unknown whether more complete use of resources at the community level correlates with resource limitation of particular species and may explain their decreasing performance with increasing plant diversity. Using the subordinate grass species Lolium perenne L. as a model, we monitored populations in 82 experimental grasslands of different plant diversity (Jena Experiment) from year 2 to 6 after establishment, and measured ecophysiological leaf traits related to light and nutrient acquisition and use. Population and plant individual sizes of L. perenne decreased with increasing species richness. A decrease in transmitted light with increasing species richness and legume proportion correlated with increasing specific leaf area (SLA). Despite this morphological adaptation to lower light availability, decreasing foliar δ(13) C signatures with increasing species richness and low variation in leaf gas exchange and chlorophyll concentrations suggested a low capacity of L. perenne for adjustment to canopy shade. Leaf nitrogen concentrations and foliar δ(15) N signatures indicated a better N supply in communities with legumes and a shift in the uptake of different N forms with increasing species richness. Leaf blade nitrate and carbohydrate concentrations as indicators of plants nutritional status supported that light limitation with increasing species richness and legume proportions, combined with a N limitation in communities with increasing proportions of non-legumes, correlated with the decreasing performance of L. perenne in communities of increasing plant diversity.

ABA inhibits germination but not dormancy release in mature imbibed seeds of Lolium rigidum Gaud.

Dormancy release in imbibed annual ryegrass (Lolium rigidum Gaud.) seeds is promoted in the dark but inhibited in the light. The role of abscisic acid (ABA) in inhibition of dormancy release was found to be negligible, compared with its subsequent effect on germination of dormant and non-dormant seeds. Inhibitors of ABA metabolism had the expected effects on seed germination but did not influence ABA concentration, suggesting that they act upon other (unknown) factors regulating dormancy. Although gibberellin (GA) synthesis was required for germination, the influence of exogenous GA on both germination and dormancy release was minor or non-existent. Embryo ABA concentration was the same following treatments to promote (dark stratification) and inhibit (light stratification) dormancy release; exogenous ABA had no effect on this process. However, the sensitivity of dark-stratified seeds to ABA supplied during germination was lower than that of light-stratified seeds. Therefore, although ABA definitely plays a role in the germination of annual ryegrass seeds, it is not the major factor mediating inhibition of dormancy release in imbibed seeds.

Green and blue light photoreceptors are involved in maintenance of dormancy in imbibed annual ryegrass (Lolium rigidum) seeds.

Light plays an important role in two separate processes within the seeds of Lolium rigidum (annual ryegrass). Dormant seeds of L. rigidum remain dormant when imbibed in the light, but once seeds have lost dormancy through dark-stratification, light stimulates their germination. This study characterizes the light qualities and quantities which are effective in maintenance of dormancy. Dormant seeds were stratified under narrow- and broad-waveband light to identify the potential photoreceptors involved in dormancy maintenance, and to determine whether dark-induced dormancy loss is reversible by light. Blue and green light both mediated dormancy maintenance in a far-red-independent manner. Red light resulted in dormancy maintenance only when far-red wavelengths were excluded, suggesting a redundant function of phytochrome. At low fluence rates, white light was more effective than monochromatic light, suggesting the action of multiple photoreceptors in dormancy maintenance. By contrast, nondormant seeds did not germinate unless provided with red light. These results indicate that seed dormancy maintenance is potentially mediated through the actions of blue and green light photoreceptors. Seed dormancy could thus be added to the growing list of plant responses that may be mediated by green light in a cryptochrome-independent manner.

Effects of ageing and microbial component on chemical availability of (137)Cs in a long-term experimental site.

A loamy soil contaminated with (137)CsCl 40 years ago was investigated by a sequential extraction technique to determine the effect of ageing on chemical availability of (137)Cs. The soil samples were sequentially extracted with H(2)O, NH(4)Ac, NH(2)OH x HCl, H(2)O(2), and HNO(3). Extractability of (137)Cs decreased in the order: HNO(3) > Residual > H(2)O(2) > NH(4)Ac > NH(2)OH x HCl > H(2)O. Only 0.94% in labile fractions (H(2)O and NH(4)Ac), while more than 96% was found in the strongly bound fraction (HNO(3) and residual). However, the activity percentage in labile fractions was increased to 1.34% after autoclaving treatment, while those in the other fractions did not significantly differ. This indicates that the microbial activity played a role in the (137)Cs retention. In the subsequent pot experiments with ryegrass and leek, specific activities in both plants were significantly higher in autoclaved soil than in non-autoclaved soil, and uptake of (137)Cs in the five cuts by ryegrass was 25% of the labile (137)Cs in the soil. In addition, a positive correlation was found between the amount of (137)Cs in labile fractions and that by plant uptake.

Flowering of the grass Lolium perenne: effects of vernalization and long days on gibberellin biosynthesis and signaling.

Almost 50 years ago, it was shown that gibberellin (GA) applications caused flowering in species normally responding to cold (vernalization) and long day (LD). The implication that GAs are involved with vernalization and LD responses is examined here with the grass Lolium perenne. This species has an obligatory requirement for exposure to both vernalization and LD for its flowering (inflorescence initiation). Specific effects of vernalization or LD on GA synthesis, content, and action have been documented using four treatment pairs: nonvernalized or vernalized plants exposed to short days (SDs) or LDs. Irrespective of vernalization status, exposure to two LDs increased expression of L. perenne GA 20-oxidase-1 (LpGA20ox1), a critical GA biosynthetic gene, with endogenous GAs increasing by up to 5-fold in leaf and shoot. In parallel, LD led to degradation of a DELLA protein, SLENDER (within 48 h of LD or within 2 h of GA application). There was no effect on GA catabolism or abscisic acid content. Loss of SLENDER, which is a repressor of GA signaling, confirms the physiological relevance of increased GA content in LD. For flowering, applied GA replaced the need for LD but not that for vernalization. Thus, GAs may be an LD, leaf-sourced hormonal signal for flowering of L. perenne. By contrast, vernalization had little impact on GA or SLENDER levels or on SLENDER degradation following GA application. Thus, although vernalization and GA are both required for flowering of L. perenne, GA signaling is independent of vernalization that apparently impacts on unrelated processes.

Photoperiodic regulation of flowering in perennial ryegrass involving a CONSTANS -like homolog.

Photoperiod and vernalization are the two key environmental factors of the floral induction of perennial ryegrass (Lolium perenne L.). Transition from vegetative to reproductive growth will only occur after an extended vernalization period, followed by an increase in day length and temperature. Here we report on the isolation and characterization of a L. perenne gene (LpCO ) that is homologous to CONSTANS , and which is tightly coupled to the floral inductive long day signal. Like other monocot CO-like proteins, the LpCO contains a zinc finger domain with a non-conserved B-Box2. Although the B-Box2 has been demonstrated to be essential for the function of the Arabidopsis CO (AtCO), LpCO is able to complement the Arabidopsis co-2 mutant, and ectopic expression in Arabidopsis wild type leads to early flowering. The LpCO transcript exhibits diurnal oscillations and is expressed at higher levels during long days.

Dormancy release during hydrated storage in Lolium rigidum seeds is dependent on temperature, light quality, and hydration status.

The influence of temperature, light environment, and seed hydration on the rate of dormancy release in Lolium rigidum (annual ryegrass) seeds during hydrated storage (stratification) was investigated. In a series of experiments, seeds were subjected to a range of temperatures (nine between 5 degrees C and 37 degrees C), light (white, red, far-red, and dark), and hydration (4-70 g H(2)O 100 g(-1) FW) during stratification for up to 80 d. Samples were germinated periodically at 25/15 degrees C or constant 15, 20, or 25 degrees C with a 12 h photoperiod to determine dormancy status. Dark-stratification was an alternative, but not equivalent dormancy release mechanism to dry after-ripening in annual ryegrass seeds. Dormancy release during dark-stratification caused a gradual increase in sensitivity to light, but germination in darkness remained negligible. Germination, but not dormancy release, was greater under fluctuating diurnal temperatures than the respective mean temperatures delivered constantly. Dormancy release rate was a positive linear function of dark-stratification temperature above a base temperature for dormancy release of 6.9 degrees C. Dormancy release at temperatures up to 30 degrees C could be described in terms of thermal dark-stratification time, but the rate of dormancy release was slower at < or =15 degrees C (244 degrees Cd/probit increase in germination) than > or =20 degrees C (208 degrees Cd/probit). Stratification in red or white, but not far-red light, inhibited dormancy release, as did insufficient (<40 g H(2)O 100 g(-1) FW) seed hydration. The influence of dark-stratification on dormancy status in annual ryegrass seeds is discussed in terms of a hypothetical increase in available membrane-bound phytochrome receptors.