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High seed quality is key to agricultural production, which is increasingly affected by climate change. We studied the effects of drought and elevated temperature during seed production on key seed quality traits of two genotypes of malting barley ( Hordeum sativum L.). Plants of a "Hana-type" landrace (B1) were taller, flowered earlier and produced heavier, larger and more vigorous seeds that resisted ageing longer compared to a semi-dwarf breeding line (B2). Accordingly, a NAC domain-containing transcription factor (TF) associated with rapid response to environmental stimuli, and the TF ABI5, a key regulator of seed dormancy and vigour, were more abundant in B1 seeds. Drought significantly reduced seed yield in both genotypes, and elevated temperature reduced seed size. Genotype B2 showed partial thermodormancy that was alleviated by drought and elevated temperature. Metabolite profiling revealed clear differences between the embryos of B1 and B2. Drought, but not elevated temperature, affected the metabolism of amino acids, organic acids, osmolytes and nitrogen assimilation, in the seeds of both genotypes. Our study may support future breeding efforts to produce new lodging and drought resistant malting barleys without trade-offs that can occur in semi-dwarf varieties such as lower stress resistance and higher dormancy., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Nagel, Arc, Rajjou, Cueff, Bailly, Clément, Sanchez-Vicente, Bailly, Seal, Roach, Rolletschek, Lorenzo, Börner and Kranner.)

Functional and structural adjustments of plants in response to environmental factors, including those occurring in alpine habitats, can result in transient acclimation, plastic phenotypic adjustments and/or heritable adaptation. To unravel repeatedly selected traits with potential adaptive advantage, we studied parallel (ecotypic) and non-parallel (regional) differentiation in leaf traits in alpine and foothill ecotypes of Arabidopsis arenosa . Leaves of plants from eight alpine and eight foothill populations, representing three independent alpine colonization events in different mountain ranges, were investigated by microscopy techniques after reciprocal transplantation. Most traits clearly differed between the foothill and the alpine ecotype, with plastic adjustments to the local environment. In alpine populations, leaves were thicker, with altered proportions of palisade and spongy parenchyma, and had fewer trichomes, and chloroplasts contained large starch grains with less stacked grana thylakoids compared to foothill populations. Geographical origin had no impact on most traits except for trichome and stomatal density on abaxial leaf surfaces. The strong parallel, heritable ecotypic differentiation in various leaf traits and the absence of regional effects suggests that most of the observed leaf traits are adaptive. These trait shifts may reflect general trends in the adaptation of leaf anatomy associated with the colonization of alpine habitats.

Background: Needle rust caused by the fungus Chrysomyxa rhododendri causes significant growth decline and increased mortality of young Norway spruce trees in subalpine forests. Extremely rare trees with enhanced resistance represent promising candidates for practice-oriented reproduction approaches. They also enable the investigation of tree molecular defence and resistance mechanisms against this fungal disease. Here, we combined RNA-Seq, RT-qPCR and secondary metabolite analyses during a period of 38 days following natural infection to investigate differences in constitutive and infection-induced defence between the resistant genotype PRA-R and three susceptible genotypes., Results: Gene expression and secondary metabolites significantly differed among genotypes from day 7 on and revealed already known, but also novel candidate genes involved in spruce molecular defence against this pathogen. Several key genes related to (here and previously identified) spruce defence pathways to needle rust were differentially expressed in PRA-R compared to susceptible genotypes, both constitutively (in non-symptomatic needles) and infection-induced (in symptomatic needles). These genes encoded both new and well-known antifungal proteins such as endochitinases and chitinases. Specific genetic characteristics concurred with varying phenolic, terpene, and hormone needle contents in the resistant genotype, among them higher accumulation of several flavonoids (mainly kaempferol and taxifolin), stilbenes, geranyl acetone, α-ionone, abscisic acid and salicylic acid., Conclusions: Combined transcriptional and metabolic profiling of the Norway spruce defence response to infection by C. rhododendri in adult trees under subalpine conditions confirmed the results previously gained on artificially infected young clones in the greenhouse, both regarding timing and development of infection, and providing new insights into genes and metabolic pathways involved. The comparison of genotypes with different degrees of susceptibility proved that several of the identified key genes are differently regulated in PRA-R, and that the resistant genotype combines a strong constitutive defence with an induced response in infected symptomatic needles following fungal invasion. Genetic and metabolic differences between the resistant and susceptible genotypes indicated a more effective hypersensitive response (HR) in needles of PRA-R that prevents penetration and spread of the rust fungus and leads to a lower proportion of symptomatic needles as well as reduced symptom development on the few affected needles., (© 2022. The Author(s).)

Trebouxiophyceae are microalgae occupying even extreme environments such as polar regions or deserts, terrestrial or aquatic, and can occur free-living or as lichen photobionts. Yet, it is poorly understood how environmental factors shape their metabolism. Here, we report on responses to light and temperature, and metabolic adjustments to desiccation in Diplosphaera epiphytica , isolated from a lichen, and Edaphochlorella mirabilis , isolated from Tundra soil, assessed via growth and photosynthetic performance parameters. Metabolite profiling was conducted by GC-MS. A meta-analysis together with data from a terrestrial and an aquatic Chlorella vulgaris strain reflected elements of phylogenetic relationship, lifestyle, and relative desiccation tolerance of the four algal strains. For example, compatible solutes associated with desiccation tolerance were up-accumulated in D. epiphytica , but also sugars and sugar alcohols typically produced by lichen photobionts. The aquatic C. vulgaris , the most desiccation-sensitive strain, showed the greatest variation in metabolite accumulation after desiccation and rehydration, whereas the most desiccation-tolerant strain, D. epiphytica , showed the least, suggesting that it has a more efficient constitutive protection from desiccation and/or that desiccation disturbed the metabolic steady-state less than in the other three strains. The authors hope that this study will stimulate more research into desiccation tolerance mechanisms in these under-investigated microorganisms.

During desiccation, the cytoplasm of orthodox seeds solidifies into an intracellular glass with highly restricted diffusion and molecular mobility. Temperature and water content govern seed ageing rates, while oxygen (O2) can promote deteriorative reactions. However, whether the cytoplasmic physical state affects involvement of O2 in seed ageing remains unresolved. We aged Pinus densiflora seeds by controlled deterioration (CD) at 45 °C and distinct relative humidity (RH), resulting in cells with a glassy (11% and 30% RH) or fluid (60% and 80% RH) cytoplasm. Hypoxic conditions (0.4% O2) during CD delayed seed deterioration, lipid peroxidation, and decline of antioxidants (glutathione, α-tocopherol, and γ-tocopherol), but only when the cytoplasm was glassy. In contrast, when the cytoplasm was fluid, seeds deteriorated at the same rate regardless of O2 availability, while being associated with limited lipid peroxidation, detoxification of lipid peroxide products, substantial loss of glutathione, and resumption of glutathione synthesis. Changes in metabolite profiles provided evidence of other O2-independent enzymatic reactions in a fluid cytoplasm, including aldo-keto reductase and glutamate decarboxylase activities. Biochemical profiles of seeds stored under seed bank conditions resembled those obtained after CD regimes that maintained a glassy cytoplasm. Overall, O2 contributed more to seed ageing when the cytoplasm was glassy, rather than fluid., (© The Author(s) 2022. Published by Oxford University Press on behalf of the Society for Experimental Biology. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

Fumarylacetoacetate hydrolase (FAH) proteins form a superfamily found in Archaea, Bacteria, and Eukaryota. However, few fumarylacetoacetate hydrolase domain (FAHD)-containing proteins have been studied in Metazoa and their role in plants remains elusive. Sequence alignments revealed high homology between two Arabidopsis thaliana FAHD-containing proteins and human FAHD1 (hFAHD1) implicated in mitochondrial dysfunction-associated senescence. Transcripts of the closest hFAHD1 orthologue in Arabidopsis (AtFAHD1a) peak during seed maturation drying, which influences seed longevity and dormancy. Here, a homology study was conducted to assess if AtFAHD1a contributes to seed longevity and vigour. We found that an A. thaliana T-DNA insertional line ( Atfahd1a-1 ) had extended seed longevity and shallower thermo-dormancy. Compared to the wild type, metabolite profiling of dry Atfahd1a-1 seeds showed that the concentrations of several amino acids, some reducing monosaccharides, and δ-tocopherol dropped, whereas the concentrations of dehydroascorbate, its catabolic intermediate threonic acid, and ascorbate accumulated. Furthermore, the redox state of the glutathione disulphide/glutathione couple shifted towards a more reducing state in dry mature Atfahd1a-1 seeds, suggesting that AtFAHD1a affects antioxidant redox poise during seed development. In summary, AtFAHD1a appears to be involved in seed redox regulation and to affect seed quality traits such as seed thermo-dormancy and longevity.

Lichens can withstand extreme desiccation to water contents of ≤ 0.1 g H2O g-1 DW, and in the desiccated state are among the most extremotolerant organisms known. Desiccation-tolerant life-forms such as seeds, mosses and lichens survive 'vitrification', that is the transition of their cytoplasm to a 'glassy' state, which causes metabolism to cease. However, our understanding of the mechanisms of desiccation tolerance is hindered by poor knowledge of what reactions occur in the desiccated state. Using Flavoparmelia caperata as a model lichen, we determined at what water contents vitrification occurred upon desiccation. Molecular mobility was assessed by dynamic mechanical thermal analysis, and the de- and re-epoxidation of the xanthophyll cycle pigments (measured by HPLC) was used as a proxy to assess enzyme activity. At 20 °C vitrification occurred between 0.12-0.08 g H2O g-1 DW and enzymes were active in a 'rubbery' state (0.17 g H2O g-1 DW) but not in a glassy state (0.03 g H2O g-1 DW). Therefore, desiccated tissues may appear to be 'dry' in the conventional sense, but subtle differences in water content will have substantial consequences on the types of (bio)chemical reactions that can occur, with downstream effects on longevity in the desiccated state., (© The Author(s) 2020. Published by Oxford University Press on behalf of the Society for Experimental Biology. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

The globally distributed green microalga Chlorella vulgaris (Chlorophyta) colonizes aquatic and terrestrial habitats, but the molecular mechanisms underpinning survival in these two contrasting environments are far from understood. Here, we compared the authentic strain of C. vulgaris from an aquatic habitat with a strain from a terrestrial high alpine habitat previously determined as Chlorella mirabilis . Molecular phylogeny of SSU rDNA (823 bp) showed that the two strains differed by one nucleotide only. Sequencing of the ITS2 region confirmed that both strains belong to the same species, but to distinct ribotypes. Therefore, the terrestrial strain was re-assessed as C. vulgaris . To study the response to environmental conditions experienced on land, we assessed the effects of irradiance and temperature on growth, of temperature on photosynthesis and respiration, and of desiccation and rehydration on photosynthetic performance. In contrast to the aquatic strain, the terrestrial strain tolerated higher temperatures and light conditions, had a higher photosynthesis-to-respiration ratio at 25°C, still grew at 30°C and was able to fully recover photosynthetic performance after desiccation at 84% relative humidity. The two strains differed most in their response to the dehydration/rehydration treatment, which was further investigated by untargeted GC-MS-based metabolite profiling to gain insights into metabolic traits differentiating the two strains. The two strains differed in their allocation of carbon and nitrogen into their primary metabolites. Overall, the terrestrial strain had higher contents of readily available nitrogen-based metabolites, especially amino acids and the polyamine putrescine. Dehydration and rehydration led to differential regulation of the amino acid metabolism, the tricarboxylic acid cycle and sucrose metabolism. The data are discussed with a view to differences in phenotypic plasticity of the two strains, and we suggest that the two genetically almost identical C. vulgaris strains are attractive models to study mechanisms that protect from abiotic stress factors, which are more frequent in terrestrial than aquatic habitats, such as desiccation and irradiation., (Copyright © 2020 Aigner, Glaser, Arc, Holzinger, Schletter, Karsten and Kranner.)

In streptophytic green algae in the genus Zygnema, pre-akinete formation is considered a key survival strategy under extreme environmental conditions in alpine and polar regions. The transition from young, dividing cells to pre-akinetes is associated with morphological changes and the accumulation of storage products. Understanding the underlying metabolic changes could provide insights into survival strategies in polar habitats. Here, GC-MS-based metabolite profiling was used to study the metabolic signature associated with pre-akinete formation in Zygnema sp. from polar regions under laboratory conditions, induced by water and nutrient depletion, or collected in the field. Light microscopy and TEM revealed drastic changes in chloroplast morphology and ultrastructure, degradation of starch grains, and accumulation of lipid bodies in pre-akinetes. Accordingly, the metabolite profiles upon pre-akinete formation reflected a gradual shift in metabolic activity. Compared with young cells, pre-akinetes showed an overall reduction in primary metabolites such as amino acids and intermediates of the tricarboxylic acid (TCA) cycle, consistent with a lower metabolic turnover, while they accumulated lipids and oligosaccharides. Overall, the transition to the pre-akinete stage involves re-allocation of photosynthetically fixed energy into storage instead of growth, supporting survival of extreme environmental conditions., (© The Author(s) 2020. Published by Oxford University Press on behalf of the Society for Experimental Biology.)

Although rice is a key crop species, few studies have addressed both rice seed physiological and nutritional quality, especially at the tissue level. In this study, an exhaustive "multi-omics" dataset on the mature rice seed was obtained by combining transcriptomics, label-free shotgun proteomics and metabolomics from embryo and endosperm, independently. These high-throughput analyses provide a new insight on the tissue-specificity related to rice seed quality. Foremost, we pinpointed that extensive post-transcriptional regulations occur at the end of rice seed development such that the embryo proteome becomes much more diversified than the endosperm proteome. Secondly, we observed that survival in the dry state in each seed compartment depends on contrasted metabolic and enzymatic apparatus in the embryo and the endosperm, respectively. Thirdly, it was remarkable to identify two different sets of starch biosynthesis enzymes as well as seed storage proteins (glutelins) in both embryo and endosperm consistently with the supernumerary embryo hypothesis origin of the endosperm. The presence of a putative new glutelin with a possible embryonic favored abundance is described here for the first time. Finally, we quantified the rate of mRNA translation into proteins. Consistently, the embryonic panel of protein translation initiation factors is much more diverse than that of the endosperm. This work emphasizes the value of tissue-specificity-centered "multi-omics" study in the seed to highlight new features even from well-characterized pathways. It paves the way for future studies of critical genetic determinants of rice seed physiological and nutritional quality.

Filamentous green algae of the genus Zygnema (Zygnematophyceae, Streptophyta) are key components of polar hydro-terrestrial mats where they face various stressors including UV irradiation, freezing, desiccation and osmotic stress. Their vegetative cells can develop into pre-akinetes, i.e. reserve-rich, mature cells. We investigated lipid accumulation and fatty acid (FA) composition upon pre-akinete formation in an Arctic and an Antarctic Zygnema strain using transmission electron microscopy and gas chromatography coupled with mass spectrometry. Pre-akinetes formed after 9 weeks of cultivation in nitrogen-free medium, which was accompanied by massive accumulation of lipid bodies. The composition of FAs was similar in both strains, and α-linolenic acid (C18:3) dominated in young vegetative cells. Pre-akinete formation coincided with a significant change in FA composition. Oleic (C18:1) and linoleic (C18:2) acid increased the most (up to 17- and 8-fold, respectively). Small amounts of long-chain polyunsaturated FAs were also detected, e.g. arachidonic (C20:4) and eicosapentaenoic (C20:5) acid. Pre-akinetes exposed to desiccation at 86% relative humidity were able to recover maximum quantum yield of photosystem II, but desiccation had no major effect on FA composition. The results are discussed with regard to the capability of Zygnema spp. to thrive in extreme conditions., (© FEMS 2016.)

Background: Lipophilic antioxidants play dual key roles in edible seeds (i) as preservatives of cell integrity and seed viability by preventing the oxidation of fats, and (ii) as essential nutrients for human and animal life stock. It has been well documented that plant domestication and post-domestication evolution frequently resulted in increased seed size and palatability, and reduced seed dormancy. Nevertheless, and surprisingly, it is poorly understood how agricultural selection and cultivation affected the physiological fitness and the nutritional quality of seeds. Fabaceae have the greatest number of crop species of all plant families, and most of them are cultivated for their highly nutritious edible seeds. Here, we evaluate whether evolution of plants under cultivation has altered the integrated system formed by membranes (fatty acids) and lipophilic antioxidants (carotenoids and tocopherols), in the ten most economically important grain legumes and their closest wild relatives, i.e.: Arachis (peanut), Cicer (chickpea), Glycine (soybean), Lathyrus(vetch), Lens (lentil), Lupinus (lupin), Phaseolus (bean), Pisum (pea), Vicia (faba bean) and Vigna (cowpea)., Results: Unexpectedly, we found that following domestication, the contents of carotenoids, including lutein and zeaxanthin, decreased in all ten species (total carotenoid content decreased 48% in average). Furthermore, the composition of carotenoids changed, whereby some carotenoids were lost in most of the crops. An undirected change in the contents of tocopherols and fatty acids was found, with contents increasing in some species and decreasing in others, independently of the changes in carotenoids. In some species, polyunsaturated fatty acids (linolenic acid especially), α-tocopherol and γ-tocopherol decreased following domestication., Conclusions: The changes in carotenoids, tocopherols and fatty acids are likely side-effects of the selection for other desired traits such as the loss of seed dormancy and dispersal mechanisms, and selection for seed storability and taste. This work may serve as baseline to broaden our knowledge on the integrated changes on crop fitness and nutritional quality following domestication.

During seed germination, the transition from a quiescent metabolic state in a dry mature seed to a proliferative metabolic state in a vigorous seedling is crucial for plant propagation as well as for optimizing crop yield. This work provides a detailed description of the dynamics of protein synthesis during the time course of germination, demonstrating that mRNA translation is both sequential and selective during this process. The complete inhibition of the germination process in the presence of the translation inhibitor cycloheximide established that mRNA translation is critical for Arabidopsis seed germination. However, the dynamics of protein turnover and the selectivity of protein synthesis (mRNA translation) during Arabidopsis seed germination have not been addressed yet. Based on our detailed knowledge of the Arabidopsis seed proteome, we have deepened our understanding of seed mRNA translation during germination by combining two-dimensional gel-based proteomics with dynamic radiolabeled proteomics using a radiolabeled amino acid precursor, namely [(35)S]-methionine, in order to highlight de novo protein synthesis, stability, and turnover. Our data confirm that during early imbibition, the Arabidopsis translatome keeps reflecting an embryonic maturation program until a certain developmental checkpoint. Furthermore, by dividing the seed germination time lapse into discrete time windows, we highlight precise and specific patterns of protein synthesis. These data refine and deepen our knowledge of the three classical phases of seed germination based on seed water uptake during imbibition and reveal that selective mRNA translation is a key feature of seed germination. Beyond the quantitative control of translational activity, both the selectivity of mRNA translation and protein turnover appear as specific regulatory systems, critical for timing the molecular events leading to successful germination and seedling establishment.

Germination ability is regulated by a combination of environmental and endogenous signals with both synergistic and antagonistic effects. Nitric oxide (NO) is a potent dormancy-releasing agent in many species, including Arabidopsis, and has been suggested to behave as an endogenous regulator of this physiological blockage. Distinct reports have also highlighted a positive impact of NO on seed germination under sub-optimal conditions. However, its molecular mode of action in the context of seed biology remains poorly documented. This review aims to focus on the implications of this radical in the control of seed dormancy and germination. The consequences of NO chemistry on the investigations on both its signaling and its targets in seeds are discussed. NO-dependent protein post-translational modifications are proposed as a key mechanism underlying NO signaling during early seed germination.

Dormancy is an adaptive trait that enables seed germination to coincide with favorable environmental conditions. It has been clearly demonstrated that dormancy is induced by abscisic acid (ABA) during seed development on the mother plant. After seed dispersal, germination is preceded by a decline in ABA in imbibed seeds, which results from ABA catabolism through 8'-hydroxylation. The hormonal balance between ABA and gibberellins (GAs) has been shown to act as an integrator of environmental cues to maintain dormancy or activate germination. The interplay of ABA with other endogenous signals is however less documented. In numerous species, ethylene counteracts ABA signaling pathways and induces germination. In Brassicaceae seeds, ethylene prevents the inhibitory effects of ABA on endosperm cap weakening, thereby facilitating endosperm rupture and radicle emergence. Moreover, enhanced seed dormancy in Arabidopsis ethylene-insensitive mutants results from greater ABA sensitivity. Conversely, ABA limits ethylene action by down-regulating its biosynthesis. Nitric oxide (NO) has been proposed as a common actor in the ABA and ethylene crosstalk in seed. Indeed, convergent evidence indicates that NO is produced rapidly after seed imbibition and promotes germination by inducing the expression of the ABA 8'-hydroxylase gene, CYP707A2, and stimulating ethylene production. The role of NO and other nitrogen-containing compounds, such as nitrate, in seed dormancy breakage and germination stimulation has been reported in several species. This review will describe our current knowledge of ABA crosstalk with ethylene and NO, both volatile compounds that have been shown to counteract ABA action in seeds and to improve dormancy release and germination.

Hydrogen peroxide (H(2)O(2)) and nitric oxide (*NO) are key reactive species in signal transduction pathways leading to activation of plant defense against biotic or abiotic stress. Here, we investigated the effect of pre-treating citrus plants (Citrus aurantium L.) with either of these two molecules on plant acclimation to salinity and show that both pre-treatments strongly reduced the detrimental phenotypical and physiological effects accompanying this stress. A proteomic analysis disclosed 85 leaf proteins that underwent significant quantitative variations in plants directly exposed to salt stress. A large part of these changes was not observed with salt-stressed plants pre-treated with either H(2)O(2) or sodium nitroprusside (SNP; a *NO-releasing chemical). We also identified several proteins undergoing changes either in their oxidation (carbonylation; 40 proteins) and/or S-nitrosylation (49 proteins) status in response to salinity stress. Both H(2)O(2) and SNP pre-treatments before salinity stress alleviated salinity-induced protein carbonylation and shifted the accumulation levels of leaf S-nitrosylated proteins to those of unstressed control plants. Altogether, the results indicate an overlap between H(2)O(2)- and *NO-signaling pathways in acclimation to salinity and suggest that the oxidation and S-nitrosylation patterns of leaf proteins are specific molecular signatures of citrus plant vigour under stressful conditions.