Plant primary carbohydrate metabolism is complex and flexible, and is regulated at many levels. Changes of transcript levels do not always lead to changes in enzyme activities, and these do not always affect metabolite levels and fluxes. To analyze interactions between these three levels of function, we have performed parallel genetic analyses of 15 enzyme activities involved in primary carbohydrate metabolism, transcript levels for their encoding structural genes, and a set of relevant metabolites. Quantitative analyses of each trait were performed in the Arabidopsis thaliana Ler x Cvi recombinant inbred line (RIL) population and subjected to correlation and quantitative trait locus (QTL) analysis.

Phytoplasmas are bacteria without cell walls from the class Mollicutes. They are obligate intracellular plant pathogens which cause diseases in hundreds of economically important plants including the grapevine (Vitis vinifera). Knowledge of their biology and the mechanisms of their interactions with hosts is largely unknown because they are uncultivable and experimentally inaccessible in their hosts. We detail here the global transcriptional profiling in grapevine responses to phytoplasmas. The gene expression patterns were followed in leaf midribs of grapevine cv. 'Chardonnay' naturally infected with a phytoplasma from the stolbur group 16SrXII-A, which is associated with the grapevine yellows disease 'Bois noir'.

Most molecular studies of plant stress tolerance have been performed with Arabidopsis thaliana, although it is not particularly stress tolerant and may lack protective mechanisms required to survive extreme environmental conditions. Thellungiella salsuginea has attracted interest as an alternative plant model species with high tolerance of various abiotic stresses. While the T. salsuginea genome has recently been sequenced, its annotation is still incomplete and transcriptomic information is scarce. In addition, functional genomics investigations in this species are severely hampered by a lack of affordable tools for genome-wide gene expression studies.

To investigate the natural variability of leaf metabolism and enzymatic activity in a maize inbred population, statistical and network analyses were employed on metabolite and enzyme profiles. The test of coefficient of variation showed that sugars and amino acids displayed opposite trends in their variance within the population, consistently with their related enzymes. The overall higher CV values for metabolites as compared to the tested enzymes are indicative for their greater phenotypic plasticity. H(2) tests revealed galactinol (1) and asparagine (0.91) as the highest scorers among metabolites and nitrate reductase (0.73), NAD-glutamate dehydrogenase (0.52), and phosphoglucomutase (0.51) among enzymes. The overall low H(2) scores for metabolites and enzymes are suggestive for a great environmental impact or gene-environment interaction. Correlation-based network generation followed by community detection analysis, partitioned the network into three main communities and one dyad, (i) reflecting the different levels of phenotypic plasticity of the two molecular classes as observed for the CV values and (ii) highlighting the concerted changes between classes of chemically related metabolites. Community 1 is composed mainly of enzymes and specialized metabolites, community 2' is enriched in N-containing compounds and phosphorylated-intermediates. The third community contains mainly organic acids and sugars. Cross-community linkages are supported by aspartate, by the photorespiration amino acids glycine and serine, by the metabolically related GABA and putrescine, and by citrate. The latter displayed the strongest node-betweenness value (185.25) of all nodes highlighting its fundamental structural role in the connectivity of the network by linking between different communities and to the also strongly connected enzyme aldolase.

Protein post-translational modifications (PTMs) are among the fastest and earliest of plant responses to changes in the environment, making the mechanisms and dynamics of PTMs an important area of plant science. One of the most studied PTMs is protein phosphorylation. This review summarizes the use of targeted proteomics for the elucidation of the biological functioning of plant PTMs, and focuses primarily on phosphorylation. Since phosphorylated peptides have a low abundance, usually complex enrichment protocols are required for their research. Initial identification is usually performed with discovery phosphoproteomics, using high sensitivity mass spectrometers, where as many phosphopeptides are measured as possible. Once a PTM site is identified, biological characterization can be addressed with targeted proteomics. In targeted proteomics, Selected/Multiple Reaction Monitoring (S/MRM) is traditionally coupled to simple, standard protein digestion protocols, often omitting the enrichment step, and relying on triple-quadruple mass spectrometer. The use of synthetic peptides as internal standards allows accurate identification, avoiding cross-reactivity typical for some antibody based approaches. Importantly, internal standards allow absolute peptide quantitation, reported down to 0.1 femtomoles, also useful for determination of phospho-site occupancy. S/MRM is advantageous in situations where monitoring and diagnostics of peptide PTM status is needed for many samples, as it has faster sample processing times, higher throughput than other approaches, and excellent quantitation and reproducibility. Furthermore, the number of publicly available data-bases with plant PTM discovery data is growing, facilitating selection of modified peptides and design of targeted proteomics workflows. Recent instrument developments result in faster scanning times, inclusion of ion-trap instruments leading to parallel reaction monitoring- which further facilitates S/MRM experimental design. Finally, recent combination of data independent and data dependent spectra acquisition means that in addition to anticipated targeted data, spectra can now be queried for unanticipated information. The potential for future applications in plant biology is outlined.

Abnormal temperatures induce physiological and biochemical changes resulting in the loss of yield. The present study investigates the impact of the PsJN strain of Paraburkholderia phytofirmans on tomato (Lycopersicon esculentum Mill.) in response to heat stress (32°C). The results of this work showed that bacterial inoculation with P. phytofirmans strain PsJN increased tomato growth parameters such as chlorophyll content and gas exchange at both normal and high temperatures (25 and 32°C). At normal temperature (25°C), the rate of photosynthesis and the photosystem II activity increased with significant accumulations of sugars, total amino acids, proline, and malate in the bacterized tomato plants, demonstrating that the PsJN strain had a positive effect on plant growth. However, the amount of sucrose, total amino acids, proline, and malate were significantly affected in tomato leaves at 32°C compared to that at 25°C. Changes in photosynthesis and chlorophyll fluorescence showed that the bacterized tomato plants were well acclimated at 32°C. These results reinforce the current knowledge about the PsJN strain of P. phytofirmans and highlight in particular its ability to alleviate the harmful effects of high temperatures by stimulating the growth and tolerance of tomato plants.

The acetic acid bacterium Gluconobacter oxydans 621H is characterized by its exceptional ability to incompletely oxidize a great variety of carbohydrates in the periplasm. The metabolism of this α-proteobacterium has been characterized to some extent, yet little is known about its transcriptomes and related data. In this study, we applied two different RNAseq approaches. Primary transcriptomes enriched for 5'-ends of transcripts were sequenced to detect transcription start sites, which allow subsequent analysis of promoter motifs, ribosome binding sites, and 5´-UTRs. Whole transcriptomes were sequenced to identify expressed genes and operon structures.

Respiration of bulky plant organs such as fleshy fruits depends on oxygen (O2) availability and often decreases with O2 concentration to avoid anoxia, but the relationship between O2 diffusional resistance and metabolic adjustments remains unclear. Melon fruit (Cucumis melo L.) was used to study relationships between O2 availability and metabolism in fleshy fruits. Enzyme activities, primary metabolites and O2 partial pressure were quantified from the periphery to the inner fruit mesocarp, at three stages of development. Hypoxia was gradually established during fruit development, but there was no strong oxygen gradient between the outer- and the inner mesocarp. These trends were confirmed by a mathematical modeling approach combining O2 diffusion equations and O2 demand estimates of the mesocarp tissue. A multivariate analysis of metabolites, enzyme activities, O2 demand and concentration reveals that metabolite gradients and enzyme capacities observed in melon fruits reflect continuous metabolic adjustments thus ensuring a timely maturation of the mesocarp. The present results suggest that the metabolic adjustments, especially the tuning of the capacity of cytochrome c oxidase (COX) to O2-availability that occurs during growth development, contribute to optimizing the O2-demand and avoiding the establishment of an O2 gradient within the flesh.

Quantitative molecular biology remains a challenge for researchers due to inconsistent approaches for control of errors in the final results. Due to several factors that can influence the final result, quantitative analysis and interpretation of qPCR data are still not trivial. Together with the development of high-throughput qPCR platforms, there is a need for a tool allowing for robust, reliable and fast nucleic acid quantification.

Omeprazole is a selective proton pump inhibitor in humans that inhibits the H+/K+-ATPase of gastric parietal cells. Omeprazole has been recently shown to act as a plant growth regulator and enhancer of salt stress tolerance. Here, we report that omeprazole treatment in hydroponically grown maize improves nitrogen uptake and assimilation. The presence of micromolar concentrations of omeprazole in the nutrient solution alleviates the chlorosis and growth inhibition induced by low nitrogen availability. Nitrate uptake and assimilation is enhanced in omeprazole treated plants through changes in nitrate reductase activity, primary metabolism, and gene expression. Omeprazole enhances nitrate assimilation through an interaction with nitrate reductase, altering its activation state and affinity for nitrate as a substrate. Omeprazole and its targets represent a novel method for enhancing nitrogen use efficiency in plants.

At the end of the annual horticultural production cycle of greenhouse-grown crops, large quantities of residual biomass are discarded. Here, we propose a new value chain to utilize horticultural leaf biomass for the extraction of secondary metabolites. To increase the secondary metabolite content of leaves, greenhouse-grown crop plants were exposed to low-cost abiotic stress treatments after the last fruit harvest. As proof of concept, we evaluated the production of the flavonoid rutin in tomato plants subjected to nitrogen deficiency. In an interdisciplinary approach, we observed the steady accumulation of rutin in young plants under nitrogen deficiency, tested the applicability of nitrogen deficiency in a commercial-like greenhouse, developed a high efficiency extraction for rutin, and evaluated the acceptance of the proposed value chain by its key actors economically. On the basis of the positive interdisciplinary evaluation, we identified opportunities and challenges for the successful establishment of horticultural leaf biomass as a novel source for secondary metabolites.

DNA sequencing was dominated by Sanger's chain termination method until the mid-2000s, when it was progressively supplanted by new sequencing technologies that can generate much larger quantities of data in a shorter time. At the forefront of these developments, long-read sequencing technologies (third-generation sequencing) can produce reads that are several kilobases in length. This greatly improves the accuracy of genome assemblies by spanning the highly repetitive segments that cause difficulty for second-generation short-read technologies. Third-generation sequencing is especially appealing for plant genomes, which can be extremely large with long stretches of highly repetitive DNA. Until recently, the low basecalling accuracy of third-generation technologies meant that accurate genome assembly required expensive, high-coverage sequencing followed by computational analysis to correct for errors. However, today's long-read technologies are more accurate and less expensive, making them the method of choice for the assembly of complex genomes. Oxford Nanopore Technologies (ONT), a third-generation platform for the sequencing of native DNA strands, is particularly suitable for the generation of high-quality assemblies of highly repetitive plant genomes. Here we discuss the benefits of ONT, especially for the plant science community, and describe the issues that remain to be addressed when using ONT for plant genome sequencing.

The common foodstuff garlic produces the potent antibiotic defense substance allicin after tissue damage. Allicin is a redox toxin that oxidizes glutathione and cellular proteins and makes garlic a highly hostile environment for non-adapted microbes. Genomic clones from a highly allicin-resistant Pseudomonas fluorescens (PfAR-1), which was isolated from garlic, conferred allicin resistance to Pseudomonas syringae and even to Escherichia coli Resistance-conferring genes had redox-related functions and were on core fragments from three similar genomic islands identified by sequencing and in silico analysis. Transposon mutagenesis and overexpression analyses revealed the contribution of individual candidate genes to allicin resistance. Taken together, our data define a multicomponent resistance mechanism against allicin in PfAR-1, achieved through horizontal gene transfer.

Plants have evolved various acclimation responses to cope with phosphate depletion, including several changes in lipid metabolism. Thereby membrane phospholipids are dephosphorylated and can be used as an internal phosphate source, while galactolipids are incorporated into the membrane to maintain membrane functionality. Still little is known about the lipidomic and transcriptomic response of plants other than Arabidopsis thaliana upon phosphate starvation. Therefore, we employed lipidomics and transcriptomics to characterize the phosphate starvation response of lipid metabolism in tomato leaves and roots. Overall, phospholipid levels decreased and galactolipids increased during the acclimation response. In addition, an early increase of triacylglycerol was observed. Interestingly, there were major differences in the acclimation response of tomato leaves and roots: leaves mainly accumulated polyunsaturated triacylglycerol, while roots showed a massive increase in galactolipid content. In line with these results, we observed transcriptional induction of phospholipid degradation and galactolipid synthesis pathways in both analyzed tissues. In contrast, other aspects of the transcriptional response, in particular, the induction of phospholipid degradation, ER-localized fatty acid desaturation and triacylglycerol assembly differed between tomato leaves and roots. These results suggest a different modulation of degraded phospholipids toward triacylglycerols and galactolipids in phosphate-starved tomato leaves and roots. Possibly the availability and composition of acyl-CoA pools and ER-derived precursors trigger the synthesis of triacylglycerols or galactolipids. As the mechanism of triacylglycerol accumulation is poorly characterized outside of seed oil formation, these findings enhance our understanding of the phosphate starvation response and of how storage lipids accumulate under stress in vegetative tissue.

Mutations in IDH induce epigenetic and transcriptional reprogramming, differentiation bias, and susceptibility to mitochondrial inhibitors in cancer cells. Here, we first show that cell lines, PDXs, and patients with acute myeloid leukemia (AML) harboring an IDH mutation displayed an enhanced mitochondrial oxidative metabolism. Along with an increase in TCA cycle intermediates, this AML-specific metabolic behavior mechanistically occurred through the increase in electron transport chain complex I activity, mitochondrial respiration, and methylation-driven CEBPα-induced fatty acid β-oxidation of IDH1 mutant cells. While IDH1 mutant inhibitor reduced 2-HG oncometabolite and CEBPα methylation, it failed to reverse FAO and OxPHOS. These mitochondrial activities were maintained through the inhibition of Akt and enhanced activation of peroxisome proliferator-activated receptor-γ coactivator-1 PGC1α upon IDH1 mutant inhibitor. Accordingly, OxPHOS inhibitors improved anti-AML efficacy of IDH mutant inhibitors in vivo. This work provides a scientific rationale for combinatory mitochondrial-targeted therapies to treat IDH mutant AML patients, especially those unresponsive to or relapsing from IDH mutant inhibitors.

An important challenge in genome assembly is haplotype phasing, that is, to reconstruct the different haplotype sequences of an individual genome. Phasing becomes considerably more difficult with increasing ploidy, which makes polyploid phasing a notoriously hard computational problem. We present a novel genetic phasing method for plant breeding with the aim to phase two deep-sequenced parental samples with the help of a large number of progeny samples sequenced at low depth. The key ideas underlying our approach are to (i) integrate the individually weak Mendelian progeny signals with a Bayesian log-likelihood model, (ii) cluster alleles according to their likelihood of co-occurrence, and (iii) assign them to haplotypes via an interval scheduling approach. We show on two deep-sequenced parental and 193 low-depth progeny potato samples that our approach computes high-quality sparse phasings and that it scales to whole genomes.

Enhancing the dietary properties of rice is crucial to contribute to alleviating hidden hunger and non-communicable diseases in rice-consuming countries. Germination is a bioprocessing approach to increase the bioavailability of nutrients in rice. However, there is a scarce information on how germination impacts the overall nutritional profile of pigmented rice sprouts (PRS). Herein, we demonstrated that germination resulted to increase levels of certain dietary compounds, such as free phenolics and micronutrients (Ca, Na, Fe, Zn, riboflavin, and biotin). Metabolomic analysis revealed the preferential accumulation of dipeptides, GABA, and flavonoids in the germination process. Genome-wide association studies of the PRS suggested the activation of specific genes such as CHS1 and UGT genes responsible for increasing certain flavonoid compounds. Haplotype analyses showed a significant difference (P < 0.05) between alleles associated with these genes. Genetic markers associated with these flavonoids were incorporated into the random forest model, improving the accuracy of prediction of multi-nutritional properties from 89.7% to 97.7%. Deploying this knowledge to breed rice with multi-nutritional properties will be timely to address double burden nutritional challenges.

In bread wheat (Triticum aestivum L.), the simultaneous improvement of both yield and grain protein is difficult because of the strong negative relationship between these two traits. However, some genotypes deviate positively from this relationship and this has been linked to their ability to take up nitrogen (N) during the post-flowering period, regardless of their N status at flowering. The physiological and genetic determinants of post-flowering N uptake relating to N satiety are poorly understood. This study uses semi-hydroponic culture of cv. Récital under controlled conditions to explore these controls. The first objective was to record the effects of contrasting N status at flowering on post-flowering nitrate (NO₃⁻) uptake under non-limiting NO₃⁻ conditions, while following the expression of key genes involved in NO₃⁻ uptake and assimilation. We found that post-flowering NO₃⁻ uptake was strongly influenced by plant N status at flowering during the first 300-400 degree-days after flowering, overlapping with a probable regulation of nitrate uptake exerted by N demand for growth. The uptake of NO₃⁻ correlated well with the expression of the gene TaNRT2.1, coding for a root NO₃⁻ transporter, which seems to play a major role in post-flowering NO₃⁻ uptake. These results provide a useful knowledge base for future investigation of genetic variability in post-flowering N uptake and may lead to concomitant gains in both grain yield and grain protein in wheat.

Symbiosis involves responses that maintain the plant host and symbiotic partner's genetic program; yet these cues are far from elucidated. Here we describe the effects of lumichrome, a flavin identified from Rhizobium spp., applied to lotus (Lotus japonicus) and tomato (Solanum lycopersicum). Combined transcriptional and metabolite analyses suggest that both species shared common pathways that were altered in response to this application under replete, sterile conditions. These included genes involved in symbiosis, as well as transcriptional and metabolic responses related to enhanced starch accumulation and altered ethylene metabolism. Lumichrome priming also resulted in altered colonization with either Mesorhizobium loti (for lotus) or Glomus intraradices/G. mossea (for tomato). It enhanced nodule number but not nodule formation in lotus; while leading to enhanced hyphae initiation and delayed arbuscule maturation in tomato.

Cell adhesion in plants is mediated predominantly by pectins, a group of complex cell wall associated polysaccharides. An Arabidopsis mutant, friable1 (frb1), was identified through a screen of T-DNA insertion lines that exhibited defective cell adhesion. Interestingly, the frb1 plants displayed both cell and organ dissociations and also ectopic defects in organ separation. The FRB1 gene encodes a Golgi-localized, plant specific protein with only weak sequence similarities to known proteins (DUF246). Unlike other cell adhesion deficient mutants, frb1 mutants do not have reduced levels of adhesion related cell wall polymers, such as pectins. Instead, FRB1 affects the abundance of galactose- and arabinose-containing oligosaccharides in the Golgi. Furthermore, frb1 mutants displayed alteration in pectin methylesterification, cell wall associated extensins and xyloglucan microstructure. We propose that abnormal FRB1 action has pleiotropic consequences on wall architecture, affecting both the extensin and pectin matrices, with consequent changes to the biomechanical properties of the wall and middle lamella, thereby influencing cell-cell adhesion.