Heat shock exposure during early wheat grain development can reduce maximum endosperm cell number but not necessarily final grain dry mass.

Post-anthesis heat shocks, which are expected to increase in frequency under climate change, may affect wheat grain development and lead to significant decreases in grain yield. Grain development occurs in three phases, the lag-phase, the filling-phase, and maturation. The growth of the three main compartments of the grain (outer layers (OLs), endosperm, embryo) is staggered, so that heat shocks affect time- and tissue-specific growth processes differentially depending on their timing. We hypothesized that heat shocks during the lag-phase may reduce final grain size, resulting from a reduction in endosperm cell number and/or a restricted OLs growth. Plants were heated for four consecutive days during the lag-phase or the filling-phase or both phases (lag- and filling-). Heat shocks consisted in four hours a day at 38°C and 21°C for the rest of the day. Controlled plants were maintained at 21/14°C (day/night). For each temperature treatment, kinetics of whole grain and compartment masses and dimensions were measured as well as the endosperm cell number. An early heat shock reduced endosperm cell proliferation. However, the growth patterns neither of endosperm nor of OLs were modified compared to controls, resulting in no differences in final grain size. Furthermore, compared to controls, a single heat shock during the filling-phase reduced both the duration and rate of dry mass accumulation into grains, whereas two consecutive shocks reduced the duration but enhanced the rate of dry mass of accumulation, even when endosperm cell number was reduced. The mean endosperm cell size was shown to be larger after early heat shocks. All together, these results suggest a compensatory mechanism exists to regulate endosperm cell size and number. This process might be a new mechanistic target for molecular studies and would improve our understanding of post-anthesis wheat tolerance to heat-shocks.

New Growth-Related Features of Wheat Grain Pericarp Revealed by Synchrotron-Based X-ray Micro-Tomography and 3D Reconstruction.

Wheat (Triticum aestivum L.) is one of the most important crops as it provides 20% of calories and proteins to the human population. To overcome the increasing demand in wheat grain production, there is a need for a higher grain yield, and this can be achieved in particular through an increase in the grain weight. Moreover, grain shape is an important trait regarding the milling performance. Both the final grain weight and shape would benefit from a comprehensive knowledge of the morphological and anatomical determinism of wheat grain growth. Synchrotron-based phase-contrast X-ray microtomography (X-ray µCT) was used to study the 3D anatomy of the growing wheat grain during the first developmental stages. Coupled with 3D reconstruction, this method revealed changes in the grain shape and new cellular features. The study focused on a particular tissue, the pericarp, which has been hypothesized to be involved in the control of grain development. We showed considerable spatio-temporal diversity in cell shape and orientations, and in tissue porosity associated with stomata detection. These results highlight the growth-related features rarely studied in cereal grains, which may contribute significantly to the final grain weight and shape.

How does post-flowering heat impact grain growth and its determining processes in wheat?

Wheat grain yield is anticipated to suffer from the increased temperatures expected under climate change. In particular, the effects of post-anthesis temperatures on grain growth and development must be better understood in order to improve crop models. Grain growth and development involve several processes, and we hypothesized that some of the most important processes, namely grain dry biomass and water accumulation, grain volume expansion, and endosperm cell proliferation, will have different thermal sensitivity. To assess this, we established temperature-response curves of these processes for steady post-anthesis temperatures between 15 °C and 36 °C. From anthesis to maturity, grain dry mass, water mass, volume, and endosperm cell number were monitored, whilst considering grain temperature. Different sensitivities to heat of these various processes were revealed. The rate of grain dry biomass accumulation increased linearly up to 25 °C, while the reciprocal of its duration increased linearly up to at least 32 °C. In contrast, the growth rates of traits contributing to grain expansion, such as increase in grain volume and cell numbers, had higher optimum temperatures, while the reciprocal of their durations were significantly lower. These temperature-response curves can contribute to improve current crop models, and allow targeting of specific mechanisms for genetic and genomic studies.

Wheat individual grain-size variance originates from crop development and from specific genetic determinism.

Wheat grain yield is usually decomposed in the yield components: number of spikes / m2, number of grains / spike, number of grains / m2 and thousand kernel weight (TKW). These are correlated one with another due to yield component compensation. Under optimal conditions, the number of grains per m2 has been identified as the main determinant of yield. However, with increasing occurrences of post-flowering abiotic stress associated with climate change, TKW may become severely limiting and hence a target for breeding. TKW is usually studied at the plot scale as it represents the average mass of a grain. However, this view disregards the large intra-genotypic variance of individual grain mass and its effect on TKW. The aim of this study is to investigate the determinism of the variance of individual grain size. We measured yield components and individual grain size variances of two large genetic wheat panels grown in two environments. We also carried out a genome-wide association study using a dense SNPs array. We show that the variance of individual grain size partly originates from the pre-flowering components of grain yield; in particular it is driven by canopy structure via its negative correlation with the number of spikes per m2. But the variance of final grain size also has a specific genetic basis. The genome-wide analysis revealed the existence of QTL with strong effects on the variance of individual grain size, independently from the other yield components. Finally, our results reveal some interesting drivers for manipulating individual grain size variance either through canopy structure or through specific chromosomal regions.

Challenging the putative structure of mannan in wheat (Triticum aestivum) endosperm.

In wheat endosperm, mannan, is poorly documented. Nevertheless, this hemicellulosic polysaccharide might have a determinant role in wheat grain development since, in Arabidopsis thaliana, mutants with a reduced amount of mannan show an altered seed development. In order to gain knowledge about mannan in wheat, we have determined its biochemical structure in wheat endosperm where mannose content is about 0.2% (dry weight basis). We developed a method of enzymatic fingerprinting and isolated mannan-enriched fractions to decipher its fine structure. Although it is widely accepted that the class of mannan present in grass cell walls is glucomannan, our data indicate that, in wheat endosperm, this hemicellulose is only represented by short unsubstituted chains of 1,4 linked D-mannose residues and is slightly acetylated. Our study provides information regarding the interactions of mannan with other cell wall components and help to progress towards the understanding of monocot cell wall architecture and the mannan synthesis in wheat endosperm.

Use of X-ray micro computed tomography imaging to analyze the morphology of wheat grain through its development.

BACKGROUND: Wheat is one of the most important staple source in the world for human consumption, animal feed and industrial raw materials. To deal with the global and increasing population demand, enhancing crop yield by increasing the final weight of individual grain is considered as a feasible solution. Morphometric analysis of wheat grain plays an important role in tracking and understanding developmental processes by assessing potential impacts on grains properties, size and shape that are major determinants of final grain weight. X-ray micro computed tomography (µCT) is a very powerful non-invasive imaging tool that is able to acquire 3D images of an individual grain, enabling to assess the morphology of wheat grain and of its different compartments. Our objective is to quantify changes of morphology during growth stages of wheat grain from 3D µCT images. METHODS: 3D µCT images of wheat grains were acquired at various development stages ranging from 60 to 310 degree days after anthesis. We developed robust methods for the identification of outer and inner tissues within the grains, and the extraction of morphometric features using 3D µCT images. We also developed a specific workflow for the quantification of the shape of the grain crease. RESULTS: The different compartments of the grain could be semi-automatically segmented. Variations of volumes of the compartments adequately describe the different stages of grain developments. The evolution of voids within wheat grain reflects lysis of outer tissues and growth of inner tissues. The crease shape could be quantified for each grain and averaged for each stage of development, helping us understand the genesis of the grain shape. CONCLUSION: This work shows that µCT acquisitions and image processing methodologies are powerful tools to extract morphometric parameters of developing wheat grain. The results of quantitative analysis revealed remarkable features of wheat grain growth. Further work will focus on building a computational model of wheat grain growth based on real 3D imaging data.

Climate change impact and adaptation for wheat protein.

Wheat grain protein concentration is an important determinant of wheat quality for human nutrition that is often overlooked in efforts to improve crop production. We tested and applied a 32-multi-model ensemble to simulate global wheat yield and quality in a changing climate. Potential benefits of elevated atmospheric CO2 concentration by 2050 on global wheat grain and protein yield are likely to be negated by impacts from rising temperature and changes in rainfall, but with considerable disparities between regions. Grain and protein yields are expected to be lower and more variable in most low-rainfall regions, with nitrogen availability limiting growth stimulus from elevated CO2 . Introducing genotypes adapted to warmer temperatures (and also considering changes in CO2 and rainfall) could boost global wheat yield by 7% and protein yield by 2%, but grain protein concentration would be reduced by -1.1 percentage points, representing a relative change of -8.6%. Climate change adaptations that benefit grain yield are not always positive for grain quality, putting additional pressure on global wheat production.

Ferulate and lignin cross-links increase in cell walls of wheat grain outer layers during late development.

Important biological, nutritional and technological roles are attributed to cell wall polymers from cereal grains. The composition of cell walls in dry wheat grain has been well studied, however less is known about cell wall deposition and modification in the grain outer layers during grain development. In this study, the composition of cell walls in the outer layers of the wheat grain (Triticum aestivum Recital cultivar) was investigated during grain development, with a focus on cell wall phenolics. We discovered that lignification of outer layers begins earlier than previously reported and long before the grain reaches its final size. Cell wall feruloylation increased in development. However, in the late stages, the amount of ferulate releasable by mild alkaline hydrolysis was reduced as well as the yield of lignin-derived thioacidolysis monomers. These reductions indicate that new ferulate-mediated cross-linkages of cell wall polymers appeared as well as new resistant interunit bonds in lignins. The formation of these additional linkages more specifically occurred in the outer pericarp. Our results raised the possibility that stiffening of cell walls occur at late development stages in the outer pericarp and might contribute to the restriction of the grain radial growth.

Both gravistimulation onset and removal trigger an increase of cytoplasmic free calcium in statocytes of roots grown in microgravity.

Gravity is a permanent environmental signal guiding plant growth and development. Gravity sensing in plants starts with the displacement of starch-filled plastids called statoliths, ultimately leading to auxin redistribution and organ curvature. While the involvement in gravity sensing of several actors such as calcium is known, the effect of statolith displacement on calcium changes remains enigmatic. Microgravity is a unique environmental condition offering the opportunity to decipher this link. In this study, roots of Brassica napus were grown aboard the International Space Station (ISS) either in microgravity or in a centrifuge simulating Earth gravity. The impact of short simulated gravity onset and removal was measured on statolith positioning and intracellular free calcium was assessed using pyroantimonate precipitates as cytosolic calcium markers. Our findings show that a ten-minute onset or removal of gravity induces very low statolith displacement, but which is, nevertheless, associated with an increase of the number of pyroantimonate precipitates. These results highlight that a change in the cytosolic calcium distribution is triggered in absence of a significant statolith displacement.

Multimodel ensembles improve predictions of crop-environment-management interactions.

A recent innovation in assessment of climate change impact on agricultural production has been to use crop multimodel ensembles (MMEs). These studies usually find large variability between individual models but that the ensemble mean (e-mean) and median (e-median) often seem to predict quite well. However, few studies have specifically been concerned with the predictive quality of those ensemble predictors. We ask what is the predictive quality of e-mean and e-median, and how does that depend on the ensemble characteristics. Our empirical results are based on five MME studies applied to wheat, using different data sets but the same 25 crop models. We show that the ensemble predictors have quite high skill and are better than most and sometimes all individual models for most groups of environments and most response variables. Mean squared error of e-mean decreases monotonically with the size of the ensemble if models are added at random, but has a minimum at usually 2-6 models if best-fit models are added first. Our theoretical results describe the ensemble using four parameters: average bias, model effect variance, environment effect variance, and interaction variance. We show analytically that mean squared error of prediction (MSEP) of e-mean will always be smaller than MSEP averaged over models and will be less than MSEP of the best model if squared bias is less than the interaction variance. If models are added to the ensemble at random, MSEP of e-mean will decrease as the inverse of ensemble size, with a minimum equal to squared bias plus interaction variance. This minimum value is not necessarily small, and so it is important to evaluate the predictive quality of e-mean for each target population of environments. These results provide new information on the advantages of ensemble predictors, but also show their limitations.

Coexpression network and phenotypic analysis identify metabolic pathways associated with the effect of warming on grain yield components in wheat.

Wheat grains are an important source of human food but current production amounts cannot meet world needs. Environmental conditions such as high temperature (above 30°C) could affect wheat production negatively. Plants from two wheat genotypes have been subjected to two growth temperature regimes. One set has been grown at an optimum daily mean temperature of 19°C while the second set of plants has been subjected to warming at 27°C from two to 13 days after anthesis (daa). While warming did not affect mean grain number per spike, it significantly reduced other yield-related indicators such as grain width, length, volume and maximal cell numbers in the endosperm. Whole genome expression analysis identified 6,258 and 5,220 genes, respectively, whose expression was affected by temperature in the two genotypes. Co-expression analysis using WGCNA (Weighted Gene Coexpression Network Analysis) uncovered modules (groups of co-expressed genes) associated with agronomic traits. In particular, modules enriched in genes related to nutrient reservoir and endopeptidase inhibitor activities were found to be positively associated with cell numbers in the endosperm. A hypothetical model pertaining to the effects of warming on gene expression and growth in wheat grain is proposed. Under moderately high temperature conditions, network analyses suggest a negative effect of the expression of genes related to seed storage proteins and starch biosynthesis on the grain size in wheat.

Different grain-filling rates explain grain-weight differences along the wheat ear.

Thousand grain weight is one of the components determining wheat grain yield. It represents the average value of individual grain weights which depends on position within the ear and on positon within the spikelet. Our objective was to quantify the influences of individual floret anthesis date, of carpel weight at anthesis and of rate and duration of grain filling, on variation in individual final grain weight. Two bread wheat cultivars were grown in a greenhouse and their ears were sampled from anthesis through to harvest. Each ear was divided into three parts-basal, central and apical-where the two proximal grains were dissected from each of two spikelets. We analysed (i) the flowering time shift within the ear and within the spikelet; and (ii) the growth kinetics during grain filling in relation to position along the ear. For both cultivars, florets located in the central part of the ear were the first to reach anthesis followed by those in the apical part and then the basal part. Within a spikelet, the floret located nearest the rachis flowered first followed by the more distal ones. We found no significant systematic effect of flowering time-shift on final grain weight. Nevertheless, grains in the central part were heavier than the basal ones (9.75% smaller) and than the apical ones (18.25% smaller). These differences were explained mainly by differences in mean grain filling rates. Analysis of growth kinetics enabled an improved explanation of the variability of individual grain weight along the ear.

Source-to-sink transport of sugar and regulation by environmental factors.

Source-to-sink transport of sugar is one of the major determinants of plant growth and relies on the efficient and controlled distribution of sucrose (and some other sugars such as raffinose and polyols) across plant organs through the phloem. However, sugar transport through the phloem can be affected by many environmental factors that alter source/sink relationships. In this paper, we summarize current knowledge about the phloem transport mechanisms and review the effects of several abiotic (water and salt stress, mineral deficiency, CO2, light, temperature, air, and soil pollutants) and biotic (mutualistic and pathogenic microbes, viruses, aphids, and parasitic plants) factors. Concerning abiotic constraints, alteration of the distribution of sugar among sinks is often reported, with some sinks as roots favored in case of mineral deficiency. Many of these constraints impair the transport function of the phloem but the exact mechanisms are far from being completely known. Phloem integrity can be disrupted (e.g., by callose deposition) and under certain conditions, phloem transport is affected, earlier than photosynthesis. Photosynthesis inhibition could result from the increase in sugar concentration due to phloem transport decrease. Biotic interactions (aphids, fungi, viruses…) also affect crop plant productivity. Recent breakthroughs have identified some of the sugar transporters involved in these interactions on the host and pathogen sides. The different data are discussed in relation to the phloem transport pathways. When possible, the link with current knowledge on the pathways at the molecular level will be highlighted.

Down-regulation of the TaGW2 gene by RNA interference results in decreased grain size and weight in wheat.

For important food crops such as wheat and rice, grain yield depends on grain number and size. In rice (Oryza sativa), GW2 was isolated from a major quantitative trait locus for yield and encodes an E3 RING ligase that negatively regulates grain size. Wheat (Triticum aestivum) has TaGW2 homologues in the A, B, and D genomes, and polymorphisms in TaGW2-A were associated with grain width. Here, to investigate TaGW2 function, RNA interference (RNAi) was used to down-regulate TaGW2 transcript levels. Transgenic wheat lines showed significantly decreased grain size-related dimensions compared with controls. Furthermore, TaGW2 knockdown also caused a significant reduction in endosperm cell number. These results indicate that TaGW2 regulates grain size in wheat, possibly by controlling endosperm cell number. Wheat and rice GW2 genes thus seem to have divergent functions, with rice GW2 negatively regulating grain size and TaGW2 positively regulating grain size. Analysis of transcription of TaGW2 homoeologues in developing grains suggested that TaGW2-A and -D act in both the division and late grain-filling phases. Furthermore, biochemical and molecular analyses revealed that TaGW2-A is a functional E3 RING ubiquitin ligase with nucleocytoplasmic subcellular partitioning. A functional nuclear export sequence responsible for TaGW2-A export from the nucleus to the cytosol and retention in the nucleolus was identified. Therefore, these results show that TaGW2 acts in the regulation of grain size and may provide an important tool for enhancement of grain yield.

Transcriptional profile analysis of E3 ligase and hormone-related genes expressed during wheat grain development.

BACKGROUND: Wheat grains are an important source of food, stock feed and raw materials for industry, but current production levels cannot meet world needs. Elucidation of the molecular mechanisms underlying wheat grain development will contribute valuable information to improving wheat cultivation. One of the most important mechanisms implicated in plant developmental processes is the ubiquitin-proteasome system (UPS). Among the different roles of the UPS, it is clear that it is essential to hormone signaling. In particular, E3 ubiquitin ligases of the UPS have been shown to play critical roles in hormone perception and signal transduction. RESULTS: A NimbleGen microarray containing 39,179 UniGenes was used to study the kinetics of gene expression during wheat grain development from the early stages of cell division to the mid-grain filling stage. By comparing 11 consecutive time-points, 9284 differentially expressed genes were identified and annotated during this study. A comparison of the temporal profiles of these genes revealed dynamic transcript accumulation profiles with major reprogramming events that occurred during the time intervals of 80-120 and 220-240°Cdays. The list of the genes expressed differentially during these transitions were identified and annotated. Emphasis was placed on E3 ligase and hormone-related genes. In total, 173 E3 ligase coding genes and 126 hormone-related genes were differentially expressed during the cell division and grain filling stages, with each family displaying a different expression profile. CONCLUSIONS: The differential expression of genes involved in the UPS and plant hormone pathways suggests that phytohormones and UPS crosstalk might play a critical role in the wheat grain developmental process. Some E3 ligase and hormone-related genes seem to be up- or down-regulated during the early and late stages of the grain development.

Proteomic analysis of peripheral layers during wheat (Triticum aestivum L.) grain development.

Grains of hexaploid wheat, Triticum aestivum (cv. Récital), were collected at 15 stages of development, from anthesis to physiological maturity, 0-700°C days (degree days after anthesis). Two hundred and seven proteins of grain peripheral layers (inner pericarp, hyaline, testa and aleurone layer) were identified by 2-DE, MALDI-TOF MS and data mining, then were classified in 16 different functional categories. Study of the protein expression over time allowed identification of five main profiles and four distinct phases of development. Composite expression curves indicated that there was a shift from metabolic processes, translation, transcription and ATP interconversion towards storage and defence processes. Protein synthesis, protein turnover, signal transduction, membrane transport and biosynthesis of secondary metabolites were the mediating functions of this shift. A picture of the dynamic processes taking place in peripheral layers during grain development was obtained in this study. It should further help in the construction of proteome reference maps for the developing peripheral layers.

Proteomic and morphological analysis of early stages of wheat grain development.

The identification of 249 proteins in the first 2 wks of wheat grain development enabled the chronological description of the early processes of grain formation. Cell division involved expression of the enzymes and proteins of the cytoskeleton and structure, DNA repair and replication enzymes and cellular metabolism enzymes (synthesis of amino acids, cell wall initiation, carbon fixation and energy production, cofactors and vitamins) with a peak expression at 125 degrees C day (degrees day after anthesis). After the first synthesis of amino acids, protein transport mechanisms, translation signals, sugar metabolism (polymerization of protein) and stress/defence proteins were activated with stable expression between 150 and 280 degrees C day. Proteins responsible for folding and degradation, including different subunits of proteasome, were highly expressed at 195 degrees C day. Proteins associated with starch granules (GBSS type 1) were present at the beginning of grain formation and increased regularly up to 280 degrees C day. Heat shock proteins (HSP70, 80, 90) were expressed throughout the early grain development stages.

Phloem sap intricacy and interplay with aphid feeding.

Aphididae feed upon the plant sieve elements (SE), where they ingest sugars, nitrogen compounds and other nutrients. For ingestion, aphid stylets penetrate SE, and because of the high hydrostatic pressure in SE, phloem sap exudes out into the stylets. Severing stylets to sample phloem exudates (i.e. stylectomy) has been used extensively for the study of phloem contents. Alternative sampling techniques are spontaneous exudation upon wounding that only works in a few plant species, and the popular EDTA-facilitated exudation technique. These approaches have allowed fundamental advances on the understanding of phloem sap composition and sieve tube physiology, which are surveyed in this review. A more complete picture of metabolites, ions, proteins and RNAs present in phloem sap is now available, which has provided large evidence for the phloem role as a signalling network in addition to its primary role in partitioning of photo-assimilates. Thus, phloem sap sampling methods can have remarkable applications to analyse plant nutrition, physiology and defence responses. Since aphid behaviour is suspected to be affected by phloem sap quality, attempts to manipulate phloem sap content were recently undertaken based on deregulation in mutant plants of genes controlling amino acid or sugar content of phloem sap. This opens up new strategies to control aphid settlement on a plant host.

Compatible plant-aphid interactions: how aphids manipulate plant responses.

To access phloem sap, aphids have developed a furtive strategy, their stylets progressing towards sieve tubes mainly through the apoplasmic compartment. Aphid feeding requires that they overcome a number of plant responses, ranging from sieve tube occlusion and activation of phytohormone-signalling pathways to expression of anti-insect molecules. In addition to bypassing plant defences, aphids have been shown to affect plant primary metabolism, which could be a strategy to improve phloem sap composition in nutrients required for their growth. During compatible interactions, leading to successful feeding and reproduction, aphids cause alterations in their host plant, including morphological changes, modified resource allocation and various local as well as systemic symptoms. Repeated salivary secretions injected from the first probe in the epidermal tissue up to ingestion of sieve-tube sap may play a crucial role in the compatibility between the aphid and the plant.

Aphid infestation causes different changes in carbon and nitrogen allocation in alfalfa stems as well as different inhibitions of longitudinal and radial expansion.

Alfalfa (Medicago sativa) stem elongation is strongly reduced by a pea aphid (Acyrthosiphon pisum Harris) infestation. As pea aphid is a phloem feeder that does not transmit virus or toxins, assimilate withdrawal is generally considered as the main mechanism responsible for growth reduction. Using a kinematic analysis, we investigated the spatial distributions of relative elemental growth rates of control and infested alfalfa stems. The water, carbon, and nitrogen contents per unit stem length were measured along the growth zone. Deposition rates and growth-sustaining fluxes were estimated from these patterns. Severe short-term aphid infestation (200 young adults over a 24-h period) induced a strong and synchronized reduction in rates of elongation and of water and carbon deposition. Reduced nitrogen content and associated negative nitrogen deposition rates were observed in some parts of the infested stems, especially in the apex. This suggested a mobilization of nitrogen from the apical part of the growth zone, converted from a sink tissue into a source tissue by aphids. Calculation of radial growth rates suggested that aphid infestation led to a smaller reduction in radial expansion than in elongation. Together with earlier observations of long-lasting effects of a short-term infestation, this supports the hypothesis that in addition to nutrient withdrawal, a thigmomorphogenesis-like mechanism is involved in the effect of aphid infestation on stem growth.