Genetic control of root morphology in response to nitrogen across rapeseed diversity.

Rapeseed (Brassica napus L.) is an oil-containing crop of great economic value but with considerable nitrogen requirement. Breeding root systems that efficiently absorb nitrogen from the soil could be a driver to ensure genetic gains for more sustainable rapeseed production. The aim of this study is to identify genomic regions that regulate root morphology in response to nitrate availability. The natural variability offered by 300 inbred lines was screened at two experimental locations. Seedlings grew hydroponically with low or elevated nitrate levels. Fifteen traits related to biomass production and root morphology were measured. On average across the panel, a low nitrate level increased the root-to-shoot biomass ratio and the lateral root length. A large phenotypic variation was observed, along with important heritability values and genotypic effects, but low genotype-by-nitrogen interactions. Genome-wide association study and bulk segregant analysis were used to identify loci regulating phenotypic traits. The first approach nominated 319 SNPs that were combined into 80 QTLs. Three QTLs identified on the A07 and C07 chromosomes were stable across nitrate levels and/or experimental locations. The second approach involved genotyping two groups of individuals from an experimental F2 population created by crossing two accessions with contrasting lateral root lengths. These individuals were found in the tails of the phenotypic distribution. Co-localized QTLs found in both mapping approaches covered a chromosomal region on the A06 chromosome. The QTL regions contained some genes putatively involved in root organogenesis and represent selection targets for redesigning the root morphology of rapeseed.

Magnesium maintains the length of the circadian period in Arabidopsis.

The circadian clock coordinates the physiological responses of a biological system to day and night rhythms through complex loops of transcriptional/translational regulation. It can respond to external stimuli and adjust generated circadian oscillations accordingly to maintain an endogenous period close to 24 h. However, the interaction between nutritional status and circadian rhythms in plants is poorly understood. Magnesium (Mg) is essential for numerous biological processes in plants, and its homeostasis is crucial to maintain optimal development and growth. Magnesium deficiency in young Arabidopsis thaliana seedlings increased the period of circadian oscillations of the CIRCADIAN CLOCK-ASSOCIATED 1 (CCA1) promoter (pCCA1:LUC) activity and dampened their amplitude under constant light in a dose-dependent manner. Although the circadian period increase caused by Mg deficiency was light dependent, it did not depend on active photosynthesis. Mathematical modeling of the Mg input into the circadian clock reproduced the experimental increase of the circadian period and suggested that Mg is likely to affect global transcription/translation levels rather than a single component of the circadian oscillator. Upon addition of a low dose of cycloheximide to perturb translation, the circadian period increased further under Mg deficiency, which was rescued when sufficient Mg was supplied, supporting the model's prediction. These findings suggest that sufficient Mg supply is required to support proper timekeeping in plants.

Root system size and root hair length are key phenes for nitrate acquisition and biomass production across natural variation in Arabidopsis.

The role of root phenes in nitrogen (N) acquisition and biomass production was evaluated in 10 contrasting natural accessions of Arabidopsis thaliana L. Seedlings were grown on vertical agar plates with two different nitrate supplies. The low N treatment increased the root to shoot biomass ratio and promoted the proliferation of lateral roots and root hairs. The cost of a larger root system did not impact shoot biomass. Greater biomass production could be achieved through increased root length or through specific root hair characteristics. A greater number of root hairs may provide a low-resistance pathway under elevated N conditions, while root hair length may enhance root zone exploration under low N conditions. The variability of N uptake and the expression levels of genes encoding nitrate transporters were measured. A positive correlation was found between root system size and high-affinity nitrate uptake, emphasizing the benefits of an exploratory root organ in N acquisition. The expression levels of NRT1.2/NPF4.6, NRT2.2, and NRT1.5/NPF7.3 negatively correlated with some root morphological traits. Such basic knowledge in Arabidopsis demonstrates the importance of root phenes to improve N acquisition and paves the way to design eudicot ideotypes.

The oxidized cellooligosaccharides confer thermotolerance in Arabidopsis by priming ethylene via heat shock factor A2.

Global climate change, especially heatwaves and aridity, is a major threat to agricultural production and food security. This requires common efforts from the scientific community to find effective solutions to better understand and protect the plant's vulnerabilities to high temperatures. The current study demonstrates the potential of cellooligosaccharides (COS), which are native and oxidized signaling molecules released by lytic polysaccharide monooxygenases (LPMO) enzymes during cell wall degradation by microbial pathogens. The extracellular perception of COS leads to the activation of damage-triggered immunity, often protecting the plant against biotic stress. However, how these signaling molecules affect abiotic stress tolerance is poorly understood. Here, we show that native COS and oxidized COS (oxiCOS) perception increase the transcript levels of several HEAT SHOCK FACTORS (HSFs) and HEAT SHOCK PROTEINS (HSPs) genes in Arabidopsis plants. However, only oxiCOS treatment triggers ethylene priming and increases thermotolerance. Furthermore, the function of the transcription factor HSFA2 is required for these processes. Altogether, our results indicate that the perception of Damage-Associated Molecular Patterns (DAMPs) may improve tolerance to adverse abiotic conditions, like exposure to high temperatures.

A Phenotyping Method of Giant Cells from Root-Knot Nematode Feeding Sites by Confocal Microscopy Highlights a Role for CHITINASE-LIKE 1 in Arabidopsis.

Most effective nematicides for the control of root-knot nematodes are banned, which demands a better understanding of the plant-nematode interaction. Understanding how gene expression in the nematode-feeding sites relates to morphological features may assist a better characterization of the interaction. However, nematode-induced galls resulting from cell-proliferation and hypertrophy hinders such observation, which would require tissue sectioning or clearing. We demonstrate that a method based on the green auto-fluorescence produced by glutaraldehyde and the tissue-clearing properties of benzyl-alcohol/benzyl-benzoate preserves the structure of the nematode-feeding sites and the plant-nematode interface with unprecedented resolution quality. This allowed us to obtain detailed measurements of the giant cells' area in an Arabidopsis line overexpressing CHITINASE-LIKE-1 (CTL1) from optical sections by confocal microscopy, assigning a role for CTL1 and adding essential data to the scarce information of the role of gene repression in giant cells. Furthermore, subcellular structures and features of the nematodes body and tissues from thick organs formed after different biotic interactions, i.e., galls, syncytia, and nodules, were clearly distinguished without embedding or sectioning in different plant species (Arabidopsis, cucumber or Medicago). The combination of this method with molecular studies will be valuable for a better understanding of the plant-biotic interactions.

Modeling the photoperiodic entrainment of the plant circadian clock.

The circadian clock is an endogenous 24 hour rhythm that helps organisms anticipate and adapt to daily and seasonal variations in environment, such as the day/night cycle or changing temperatures. The plant clock is a complex network of transcription factors that regulate each other, forming interlocked feedback loops. Most of its components are light-regulated in some way, making the system highly sensitive to changes in light conditions. Here, we explore the mechanisms by which the plant clock adapts to changing day length. We first present some experimental data illustrating the variety of behaviors found in seedlings exposed to external day/night cycles different from 24h. We then use a mathematical model to characterize the response of the clock to a wide range of external cycle lengths and photoperiods. We show the existence of several domains of periodic entrainment with different ratios between the external cycle length and the period of the clock, and the presence of quasiperiodic and chaotic behaviors outside of the entrainment range. We simulate knockout mutants with impaired clock function and theoretical variants with diminished light sensitivity to highlight the role of a complex network and multiple light inputs in keeping the clock entrained over a wide range of conditions.

Towards the discovery of novel genetic component involved in stress resistance in Arabidopsis thaliana.

The exposure of plants to high concentrations of trace metallic elements such as copper involves a remodeling of the root system, characterized by a primary root growth inhibition and an increase in the lateral root density. These characteristics constitute easy and suitable markers for screening mutants altered in their response to copper excess. A forward genetic approach was undertaken in order to discover novel genetic factors involved in the response to copper excess. A Cu(2+) -sensitive mutant named copper modified resistance1 (cmr1) was isolated and a causative mutation in the CMR1 gene was identified by using positional cloning and next-generation sequencing. CMR1 encodes a plant-specific protein of unknown function. The analysis of the cmr1 mutant indicates that the CMR1 protein is required for optimal growth under normal conditions and has an essential role in the stress response. Impairment of the CMR1 activity alters root growth through aberrant activity of the root meristem, and modifies potassium concentration and hormonal balance (ethylene production and auxin accumulation). Our data support a putative role for CMR1 in cell division regulation and meristem maintenance. Research on the role of CMR1 will contribute to the understanding of the plasticity of plants in response to changing environments.

Targeted metabolomics and transcript profiling of methyltransferases in three coffee species.

Coffee plants contain well-known xanthines as caffeine. Three Coffea species grown in a controlled greenhouse environment were the focus of this research. Coffea arabica and C. canephora are two first principal commercial species and commonly known as arabica and robusta, respectively. Originating in Central Africa, C. anthonyi is a novel species with small leaves. The xanthine metabolites in flower, fruit and leaf extracts were compared using both targeted and untargeted metabolomics approaches. We evaluated how the xanthine derivatives and FQA isomers relate to the expression of biosynthetic genes encoding N- and O-methyltransferases. Theobromine built up in leaves of C. anthonyi because caffeine biosynthesis was hindered in the absence of synthase gene expression. Despite this, green fruits expressed these genes and they produced caffeine. Given that C. anthonyi evolved successfully over time, these findings put into question the defensive role of caffeine in leaves. An overview of the histolocalisation of xanthines in the different flower parts of Coffea arabica was also provided. The gynoecium contained more theobromine than the flower buds or petals. This could be attributed to increased caffeine biosynthesis before fructification. The presence of theophylline and the absence of theobromine in the petals indicate that caffeine is catabolized more in the petals than in the gynoecium.

Current methods for detecting ethylene in plants.

BACKGROUND: In view of ethylene's critical developmental and physiological roles the gaseous hormone remains an active research topic for plant biologists. Progress has been made to understand the ethylene biosynthesis pathway and the mechanisms of perception and action. Still numerous questions need to be answered and findings to be validated. Monitoring gas production will very often complete the picture of any ethylene research topic. Therefore the search for suitable ethylene measuring methods for various plant samples either in the field, greenhouses, laboratories or storage facilities is strongly motivated. SCOPE: This review presents an update of the current methods for ethylene monitoring in plants. It focuses on the three most-used methods - gas chromatography detection, electrochemical sensing and optical detection - and compares them in terms of sensitivity, selectivity, time response and price. Guidelines are provided for proper selection and application of the described sensor methodologies and some specific applications are illustrated of laser-based detector for monitoring ethylene given off by Arabidopsis thaliana upon various nutritional treatments. CONCLUSIONS: Each method has its advantages and limitations. The choice for the suitable ethylene sensor needs careful consideration and is driven by the requirements for a specific application.

Transcriptomic and physiological approaches to decipher cold stress mitigation exerted by brown-seaweed extract application in tomato.

Chilling temperatures represent a challenge for crop species originating from warm geographical areas. In this situation, biostimulants serve as an eco-friendly resource to mitigate cold stress in crops. Tomato (Solanum lycopersicum L.) is an economically important vegetable crop, but quite sensitive to cold stress, which it encounters in both open field and greenhouse settings. In this study, the biostimulant effect of a brown-seaweed extract (BSE) has been evaluated in tomato exposed to low temperature. To assess the product effects, physiological and molecular characterizations were conducted. Under cold stress conditions, stomatal conductance, net photosynthesis, and yield were significantly (p ≤ 0.05) higher in BSE-treated plants compared to the untreated ones. A global transcriptomic survey after BSE application revealed the impact of the BSE treatment on genes leading to key responses to cold stress. This was highlighted by the significantly enriched GO categories relative to proline (GO:0006560), flavonoids (GO:0009812, GO:0009813), and chlorophyll (GO:0015994). Molecular data were integrated by biochemical analysis showing that the BSE treatment causes greater proline, polyphenols, flavonoids, tannins, and carotenoids contents.The study highlighted the role of antioxidant molecules to enhance tomato tolerance to low temperature mediated by BSE-based biostimulant.

Dissecting the role of CHITINASE-LIKE1 in nitrate-dependent changes in root architecture.

The root phenotype of an Arabidopsis (Arabidopsis thaliana) mutant of CHITINASE-LIKE1 (CTL1), called arm (for anion-related root morphology), was previously shown to be conditional on growth on high nitrate, chloride, or sucrose. Mutants grown under restrictive conditions displayed inhibition of primary root growth, radial swelling, proliferation of lateral roots, and increased root hair density. We found here that the spatial pattern of CTL1 expression was mainly in the root and root tips during seedling development and that the protein localized to the cell wall. Fourier-transform infrared microspectroscopy of mutant root tissues indicated differences in spectra assigned to linkages in cellulose and pectin. Indeed, root cell wall polymer composition analysis revealed that the arm mutant contained less crystalline cellulose and reduced methylesterification of pectins. We also explored the implication of growth regulators on the phenotype of the mutant response to the nitrate supply. Exogenous abscisic acid application inhibited more drastically primary root growth in the arm mutant but failed to repress lateral branching compared with the wild type. Cytokinin levels were higher in the arm root, but there were no changes in mitotic activity, suggesting that cytokinin is not directly involved in the mutant phenotype. Ethylene production was higher in arm but inversely proportional to the nitrate concentration in the medium. Interestingly, eto2 and eto3 ethylene overproduction mutants mimicked some of the conditional root characteristics of the arm mutant on high nitrate. Our data suggest that ethylene may be involved in the arm mutant phenotype, albeit indirectly, rather than functioning as a primary signal.

Equations for solar tracking.

Direct sunlight absorption by trace gases can be used to quantify them and investigate atmospheric chemistry. In such experiments, the main optical apparatus is often a grating or a Fourier transform spectrometer. A solar tracker based on motorized rotating mirrors is commonly used to direct the light along the spectrometer axis, correcting for the apparent rotation of the Sun. Calculating the Sun azimuth and altitude for a given time and location can be achieved with high accuracy but different sources of angular offsets appear in practice when positioning the mirrors. A feedback on the motors, using a light position sensor close to the spectrometer, is almost always needed. This paper aims to gather the main geometrical formulas necessary for the use of a widely used kind of solar tracker, based on two 45° mirrors in altazimuthal set-up with a light sensor on the spectrometer, and to illustrate them with a tracker developed by our group for atmospheric research.

A dual-omics approach for profiling plant responses to biostimulant applications under controlled and field conditions.

A comprehensive approach using phenomics and global transcriptomics for dissecting plant response to biostimulants is illustrated with tomato (Solanum lycopersicum cv. Micro-Tom and Rio Grande) plants cultivated in the laboratory, greenhouse, and open field conditions. Biostimulant treatment based on an Ascophyllum nodosum extract (ANE) was applied as a foliar spray with two doses (1 or 2 l ha-1) at three different phenological stages (BBCH51, BBCH61, and BBCH65) during the flowering phase. Both ANE doses resulted in greater net photosynthesis rate, stomatal conductance, and fruit yield across all culture conditions. A global transcriptomic analysis of leaves from plants grown in the climate chamber, revealed a greater number of differentially expressed genes (DEGs) with the low ANE dose compared to the greater one. The second and third applications induced broader transcriptome changes compared to the first one, indicating a cumulative treatment effect. The functional enrichment analysis of DEGs highlighted pathways related to stimulus-response and photosynthesis, consistent with the morpho-physiological observations. This study is the first comprehensive dual-omics approach for profiling plant responses to biostimulants across three different culture conditions.

Low magnesium status in plants enhances tolerance to cadmium exposure.

In a transcriptomic study of magnesium (Mg) starvation in Arabidopsis, we identified several genes that were differentially regulated which are involved in the detoxification process of nonessential heavy metals such as cadmium (Cd). We further tested the impact of low Mg status on Cd sensitivity in plants. Interestingly, a -Mg pretreatment of 7 d alleviated the bleaching of young leaves caused by Cd. No or little difference in Cd tissue concentration between the +Mg and -Mg plants was observed, suggesting that lower Cd toxicity was probably not attributable to modified root to shoot translocation. Mg deficiency also promoted an increase in the iron (Fe) concentration (up to one-fourth) in Cd-treated leaves. Because high Fe concentrations have previously been reported to prevent the harmful effects of Cd, we explored whether Fe homeostasis plays a role in the Mg-Cd interaction. A protective effect of -Mg pretreatment was also observed on Fe starvation. However, Fe foliar spray partially alleviated Cd-induced chloroses, while it almost completely restored chlorophyll content in Fe-deficient leaves. In conclusion, the protective effect of Mg against Cd toxicity could be attributable partly to the maintenance of Fe status but also to the increase in antioxidative capacity, detoxification and/or protection of the photosynthetic apparatus.

Chitinase-like protein CTL1 plays a role in altering root system architecture in response to multiple environmental conditions.

Plant root architecture is highly responsive to changes in nutrient availability. However, the molecular mechanisms governing the adaptability of root systems to changing environmental conditions is poorly understood. A screen for abnormal root architecture responses to high nitrate in the growth medium was carried out for a population of ethyl methanesulfonate-mutagenized Arabidopsis (Arabidopsis thaliana). The growth and root architecture of the arm (for anion altered root morphology) mutant described here was similar to wild-type plants when grown on low to moderate nitrate concentrations, but on high nitrate, arm exhibited reduced primary root elongation, radial swelling, increased numbers of lateral roots, and increased root hair density when compared to the wild-type control. High concentrations of chloride and sucrose induced the same phenotype. In contrast, hypocotyl elongation in the dark was decreased independently of nitrate availability. Positional cloning identified a point mutation in the AtCTL1 gene that encodes a chitinase-related protein, although molecular and biochemical analysis showed that this protein does not possess chitinase enzymatic activity. CTL1 appears to play two roles in plant growth and development based on the constitutive effect of the arm mutation on primary root growth and its conditional impact on root architecture. We hypothesize that CTL1 plays a role in determining cell wall rigidity and that the activity is differentially regulated by pathways that are triggered by environmental conditions. Moreover, we show that mutants of some subunits of the cellulose synthase complex phenocopy the conditional effect on root architecture under nonpermissive conditions, suggesting they are also differentially regulated in response to a changing environment.

LPMO-oxidized cellulose oligosaccharides evoke immunity in Arabidopsis conferring resistance towards necrotrophic fungus B. cinerea.

Lytic Polysaccharide Monooxygenases (LPMOs) are powerful redox enzymes able to oxidatively cleave recalcitrant polysaccharides. Widely conserved across biological kingdoms, LPMOs of the AA9 family are deployed by phytopathogens to deconstruct cellulose polymers. In response, plants have evolved sophisticated mechanisms to sense cell wall damage and thus self-triggering Damage Triggered Immunity responses. Here, we show that Arabidopsis plants exposed to LPMO products triggered the innate immunity ultimately leading to increased resistance to the necrotrophic fungus Botrytis cinerea. We demonstrated that plants undergo a deep transcriptional reprogramming upon elicitation with AA9 derived cellulose- or cello-oligosaccharides (AA9_COS). To decipher the specific effects of native and oxidized LPMO-generated AA9_COS, a pairwise comparison with cellobiose, the smallest non-oxidized unit constituting cellulose, is presented. Moreover, we identified two leucine-rich repeat receptor-like kinases, namely STRESS INDUCED FACTOR 2 and 4, playing a crucial role in signaling the AA9_COS-dependent responses such as camalexin production. Furthermore, increased levels of ethylene, jasmonic and salicylic acid hormones, along with deposition of callose in the cell wall was observed. Collectively, our data reveal that LPMOs might play a crucial role in plant-pathogen interactions.

Early transcriptomic changes induced by magnesium deficiency in Arabidopsis thaliana reveal the alteration of circadian clock gene expression in roots and the triggering of abscisic acid-responsive genes.

*Plant growth and development ultimately depend on environmental variables such as the availability of essential minerals. Unravelling how nutrients affect gene expression will help to understand how they regulate plant growth. *This study reports the early transcriptomic response to magnesium (Mg) deprivation in Arabidopsis. Whole-genome transcriptome was studied in the roots and young mature leaves 4, 8 and 28 h after the removal of Mg from the nutrient solution. *The highest number of regulated genes was first observed in the roots. Contrary to other mineral deficiencies, Mg depletion did not induce a higher expression of annotated genes in Mg uptake. Remarkable responses include the perturbation of the central oscillator of the circadian clock in roots and the triggering of abscisic acid (ABA) signalling, with half of the up-regulated Mg genes in leaves being ABA-responsive. However, no change in ABA content was observed. *The specificity of the response of some Mg-regulated genes was challenged by studying their expression after other mineral deficiencies and environmental stresses. The possibility to develop markers for Mg incipient deficiency is discussed here.

Systems analysis of the responses to long-term magnesium deficiency and restoration in Arabidopsis thaliana.

*Unravelling mechanisms that control plant growth as a function of nutrient availability presents a major challenge in plant biology. This study reports the first transcriptome response to long-term (1 wk) magnesium (Mg) depletion and restoration in Arabidopsis thaliana. *Before the outbreak of visual symptoms, genes responding to Mg starvation and restoration were monitored in the roots and young mature leaves and compared with the Mg fully supplied as control. *After 1 wk Mg starvation in roots and leaves, 114 and 2991 genes were identified to be differentially regulated, respectively, which confirmed the later observation that the shoot development was more affected than the root in Arabidopsis. After 24 h of Mg resupply, restoration was effective for the expression of half of the genes altered. We emphasized differences in the expression amplitude of genes associated with the circadian clock predominantly in leaves, a higher expression of genes in the ethylene biosynthetic pathway, in the reactive oxygen species detoxification and in the photoprotection of the photosynthetic apparatus. Some of these observations at the molecular level were verified by metabolite analysis. *The results obtained here will help us to better understand how changes in Mg availability are translated into adaptive responses in the plant.