β-Glucosidase is validated as an elicitor for early immune responses in plants and it was detected in the salivary glands of Frankliniella occidentalis in previous research. Seven differentially expressed genes encoding β-Glucosidase were obtained by comparing the transcriptomes of F. occidentalis adults grown under two different CO 2 concentrations (800 ppm vs. 400 ppm), which might be associated with the differences in the interaction between F. occidentalis adults and its host plant, Phaseolus vulgaris under different CO 2 levels. To verify this speculation, changes in defense responses based on the production and elimination of reactive oxygen species (ROS) in P. vulgaris leaves treated with three levels of β-Glucosidase activity under ambient CO 2 (aCO 2) and elevated CO 2 (eCO 2) were measured in this study. The results showed that both leaves infested with thrips and those sprayed with the pure β-Glucosidase solution showed significant increases in ROS levels under aCO 2 and eCO 2, and the activities of antioxidant enzymes including superoxide dismutase (SOD), peroxidase (POD), and catalase (CAT) were increased correspondingly, while in leaves infested with FoβGlu-1-silenced thrips, the ROS levels and activities of these enzymes did not change significantly during the first 12 hours of injury regardless of CO 2 level. Besides, significantly higher levels of ROS and lower activities of SOD, POD and CAT in injured leaves under eCO 2 compared to aCO 2 were noticed, which would negatively affect P. vulgaris leaves and facilitate thrips damage.

Group VII Ethylene Response Factors (ERF-VII) are plant-specific transcription factors (TFs) known for their role in the activation of hypoxia-responsive genes under low oxygen stress but also in plant endogenous hypoxic niches. However, their function in the microaerophilic nitrogen-fixing nodules of legumes has not yet been investigated. We investigated regulation and the function of the two Medicago truncatula ERF-VII TFs ( MtERF74 and MtERF75) in roots and nodules, MtERF74 and MtERF75 in response to hypoxia stress and during the nodulation process using an RNA interference strategy and targeted proteolysis of MtERF75. Knockdown of MtERF74 and MtERF75 partially blocked the induction of hypoxia-responsive genes in roots exposed to hypoxia stress. In addition, a significant reduction in nodulation capacity and nitrogen fixation activity was observed in mature nodules of double knockdown transgenic roots. Overall, the results indicate that MtERF74 and MtERF75 are involved in the induction of MtNR1 and Pgb1.1 expression for efficient Phytogb-NO respiration in the nodule.

Plant metabolomics has been used widely in plant physiology, in particular to analyse metabolic responses to environmental parameters. Derivatization (via trimethylsilylation-methoximation) followed by GC-MS metabolic profiling is a major technique to quantify low molecular weight, common metabolites of primary carbon, sulphur and nitrogen metabolism. There are now excellent opportunities for new generation analyses, using high resolution, exact mass GC-MS spectrometers that are progressively becoming relatively cheap. However, exact mass GC-MS analyses for routine metabolic profiling are not common, there is no dedicated available database. Also, exact mass GC-MS is usually dedicated to structural resolution of targeted secondary metabolites. Here, we present a curated database for exact mass metabolic profiling (made of 336 analytes, 1,064 characteristic exact mass fragments) focused on molecules of primary metabolism. We show advantages of exact mass analyses, in particular to resolve isotopic patterns, localise S-containing metabolites, and avoid identification errors when analytes have common nominal mass peaks in their spectrum. We provide a practical example using leaves of different Arabidopsis ecotypes and show how exact mass GC-MS analysis can be applied to plant samples and identify metabolic profiles.

Triose-phosphate utilization (TPU) limits the maximum rate at which plants can photosynthesize. However, TPU is almost never found to be limiting photosynthesis under ambient conditions for plants. This, along with previous results showing adaptability of TPU at low temperature, suggest that TPU capacity is regulated to be just above the photosynthetic rate achievable under the prevailing conditions. A set of experiments were performed to study the adaptability of TPU capacity when plants are acclimated to elevated CO 2 concentrations. Plants held at 1500 ppm CO 2 were initially TPU limited. After 30 hours they no longer exhibited TPU limitations but they did not elevate their TPU capacity. Instead, the maximum rates of carboxylation and electron transport declined. A timecourse of regulatory responses was established. A step increase of CO 2 first caused PSI to be oxidized but after 40 s both PSI and PSII had excess electrons as a result of acceptor-side limitations. Electron flow to PSI slowed and the proton motive force increased. Eventually, non-photochemical quenching reduced electron flow sufficiently to balance the TPU limitation. Over several minutes rubisco deactivated contributing to regulation of metabolism to overcome the TPU limitation.

N-terminal cysteine oxidases (NCOs) use molecular oxygen to oxidize the amino-terminal cysteine of specific proteins, thereby initiating the proteolytic N-degron pathway. To expand the characterization of the plant family of NCOs (PCOs), we performed a phylogenetic analysis across different taxa in terms of sequence similarity and transcriptional regulation. Based on this survey, we propose a distinction of PCOs into two main groups. A-type PCOs are conserved across all plant species and are generally unaffected at the mRNA level by oxygen availability. Instead, B-type PCOs differentiated in spermatophytes to acquire transcriptional regulation in response to hypoxia. The inactivation of two A-type PCOs in Arabidopsis thaliana, PCO4 and PCO5, is sufficient to activate the anaerobic response in young seedlings, whereas the additional removal of B-type PCOs leads to a stronger induction of anaerobic genes and impairs plant growth and development. Our results show that both PCO types are required to regulate the anaerobic response in angiosperms. Therefore, while it is possible to distinguish two clades within the PCO family, we conclude that they all contribute to restrain the anaerobic transcriptional program in normoxic conditions and together generate a molecular switch to toggle the hypoxic response.

Microbe associated molecular pattern (MAMP) triggered immunity research has traditionally centred around signal transduction pathways originating from activated membrane localised pattern recognition receptors (PRRs), culminating in nuclear transcription and post translational modifications. More recently, chloroplasts have emerged as key immune signalling hubs. Chloroplasts play a central role in integrating environmental signals. Notably MAMP recognition induces chloroplastic ROS (cROS) which is suppressed by pathogens effectors, which also modify the balance of defence hormone precursors, jasmonic acid (JA), salicylic acid (SA) and abscisic acid (ABA), whose precursors are chloroplast synthesised. This study focuses on how well characterised PRRs and co-receptors modulate chloroplast physiology, examining whether diverse signalling pathways converge to similarly modulate chloroplast function. Pre-treatment of receptor mutant plants with MAMP and D(Damage)AMP peptides usually protect against effector modulation of chlorophyll fluorescence and prevent Pseudomonas syringae effector mediated quenching of cROS and suppression of Fv/Fm. The MAMP-triggered immunity (MTI) co-receptor double mutant, bak1-5/bkk1-1, exhibits a remarkable decrease in Fv/Fm compared to control plants during infection, underlining the importance of MTI mediated signalling in chloroplast immunity. Further probing the role of the chloroplast in immunity we unexpectedly found that high light uncouples plant immune signalling.

Calcium (Ca 2+) is an important second messenger in plants. The activation of Ca 2+ signaling cascades is critical in the activation of adaptive processes in response to perceived environmental stimuli, including biotic stresses. The colonization of roots by the plant growth promoting endophyte Serendipita indica involves the increase of cytosolic Ca 2+ levels in Arabidopsis thaliana. In this study, we investigated transcriptional changes in Arabidopsis roots during symbiosis with S. indica. RNA-seq profiling disclosed the significant induction of CALCINEURIN B-LIKE 7 ( CBL7) during early- and later phases of the interaction. Consistent with the transcriptomics analysis, reverse genetic evidence and yeast two-hybrid studies highlighted the functional relevance of CBL7 and tested the involvement of a CBL7-CBL-INTERACTING PROTEIN KINASE 13 (CIPK13) signaling pathway in the establishment of the mutualistic relationship that promotes plant growth. The loss-of-function of CBL7 abolished the growth promoting effect of S. indica and affected the colonization of the root by the fungus. The subsequent transcriptomics analysis of cbl7 revealed the involvement of this Ca 2+ sensor in activating plant defense responses. Furthermore, we report on the contribution of CBL7 to potassium transport in Arabidopsis. Triggered by the differential expression of a small number of K + channels/transporter genes, we analyzed K + contents in wild-type and cbl7 plants and observed a significant accumulation of K + in root of cbl7 plants, while shoot tissues demonstrated K + depletion. Taken together, our work associates CBL7 with an important role in the mutual interaction between Arabidopsis and S. indica and links the CBL7 Ca 2+ receptor protein to K + transport.

Plant vacuoles serve as the primary intracellular compartments for phosphorus (P) storage and play a central role to maintain P homeostasis. The Oryza sativa (rice) genome contains three genes that encode SPX (SYG1/PHO81/XPR1)-MFS (Major Facility Superfamily) proteins (OsSPX-MFS1, 2, 3). OsSPX-MFS1 and OsSPX-MFS3 were shown previously to have vacuolar phosphate (Pi) transporter activities, but the physiological role of the three transporters under varying P conditions and under field grown conditions for a crop plant is not known. To address this knowledge gap, we generated single, double, and triple mutants (7 mutants with at least two lines of each) for the three rice Os SPX-MFS genes. All the mutants except osspx-mfs2 display lower vacuolar Pi concentrations and all Os SPX-MFSs overexpression plant lines display higher Pi accumulation, demonstrating that all three OsSPX-MFSs are vacuolar Pi influx transporters. OsSPX-MFS3 plays the dominant role based on the phenotypes of three single mutants in terms of growth, vacuolar and tissue Pi concentrations. OsSPX-MFS2 is the weakest and only functions as vacuole Pi sequestration under osspx-mfs1/3 background. The vacuolar Pi sequestration was severely impaired in osspx-mfs1/ 3 and osspx-mfs1/2/ 3, which led to Pi toxicity and subsequently increased Pi allocation to aerial organs. High Pi in the panicle result in necrotic symptoms on husks and impaired panicle and grain development in osspx-mfs1/ 3 and osspx-mfs1/2/ 3 mutant lines. The mutation in the weak vacuolar Pi transporter OsSPX-MFS2 resulted more stable yield compared to the wildtype under low P field conditions. The results suggest that alteration of vacuolar Pi sequestration may be a novel effective strategy to improve rice (crop) tolerance to low phosphorus field conditions and maintain yield.

Recent progress has shown that vacuolar Pi transporters (VPTs) are important for cellular Pi homeostasis against external Pi variations in Arabidopsis and rice, while it is poorly understood for the identity and regulatory mechanism of VPTs in Brassica napus ( B. napus). Here, we identified two vacuolar Pi influx transporters BnA09PHT5;1b and BnCnPHT5;1b in B. napus and uncovered their necessity for cellular Pi homeostasis through functional analysis. BnPHT5;1bs are the homologs of Arabidopsis AtPHT5;1 with the similar sequence, structure, tonoplast localization, and VPT activity. BnPHT5;1b double mutants had smaller shoot growth and higher shoot cellular Pi than the wild-type B. napus, which are largely different from the report in At PHT5;1 mutant, suggesting PHT5;1-VPTs play a distinct mechanism of cellular Pi homeostasis in seedlings of B. napus and Arabidopsis. By contrast, disruption of BnPHT5;1b genes slowed vegetative growth accompanied by Pi toxicity in floral organs, reduced seed yield and impacted seed traits, agreeing with the role of AtPHT5;1 in floral Pi homeostasis. Taken together, our studies identified two vacuolar Pi influx transporters in B. napus and revealed the distinct and conserved regulatory mechanisms of BnPHT5;1bs in cellular Pi homeostasis in this plant species.

Various root-colonizing bacterial species can promote plant growth and trigger systemic resistance against aboveground leaf pathogens and herbivore insects. To date, the underlying metabolic signatures of these rhizobacteria-induced plant phenotypes are poorly understood. To identify core metabolic pathways that are targeted by growth-promoting rhizobacteria, we used combinations of three plant species and three rhizobacterial species and interrogated plant shoot chemistry by untargeted metabolomics. A substantial part (50-64%) of the metabolites detected in plant shoot tissue was differentially affected by the rhizobacteria. Among others, the phenylpropanoid pathway was targeted by the rhizobacteria in each of the three plant species. Differential regulation of the various branches of the phenylpropanoid pathways showed an association with either plant growth promotion or growth reduction. Overall, suppression of flavonoid biosynthesis was associated with growth promotion, while growth reduction showed elevated levels of flavonoids. Subsequent assays with twelve Arabidopsis flavonoid biosynthetic mutants revealed that the proanthocyanidin branch plays an essential role in rhizobacteria-mediated growth promotion. Our study also showed that a number of pharmaceutically and nutritionally relevant metabolites in the plant shoot were significantly increased by rhizobacterial treatment, providing new avenues to use rhizobacteria to tilt plant metabolism towards the biosynthesis of valuable natural plant products.

Water inside plants forms a continuous chain from water in soils to the water evaporating from leaf surfaces. Failures in this chain result in reduced transpiration and photosynthesis and these failures are caused by soil drying and/or cavitation-induced xylem embolism. Xylem embolism and plant hydraulic failure share a number of analogies to “catastrophe theory” in dynamical systems. These catastrophes are often represented in the physiological and ecological literature as tipping points or alternative stable states when control variables exogenous (e.g. soil water potential) or endogenous (e.g. leaf water potential) to the plant are allowed to slowly vary. Here, plant hydraulics viewed from the perspective of catastrophes at multiple spatial scales is considered with attention to bubble expansion (i.e. cavitation), organ-scale vulnerability to embolism, and whole-plant biomass as a proxy for transpiration and hydraulic function. The hydraulic safety-efficiency tradeoff, hydraulic segmentation and maximum plant transpiration are examined using this framework. Underlying mechanisms for hydraulic failure at very fine scales such as pit membranes, intermediate scales such as xylem network properties and at larger scales such as soil-tree hydraulic pathways are discussed. Lacunarity areas in plant hydraulics are also flagged where progress is urgently needed.

The coordination of plant leaf water potential (ΨL) regulation and xylem vulnerability to embolism is fundamental for understanding the tradeoffs between carbon uptake and risk of hydraulic damage. There is a general consensus that trees with vulnerable xylem regulate ΨL more conservatively than plants with resistant xylem. We evaluated if this paradigm applied to three important eastern US temperate tree species, Quercus alba L., Acer saccharum Marsh., and Liriodendron tulipifera L., by synthesizing 1600 ΨL observations, 122 xylem embolism curves, and xylem anatomical measurements across ten forests spanning pronounced hydroclimatological gradients and ages. We found that, unexpectedly, the species with the most vulnerable xylem (Q. alba) regulated ΨL less strictly than the other species. This relationship was found across all sites, such that coordination among traits was largely unaffected by climate and stand age. Quercus species are perceived to be among the most drought tolerant temperate US forest species; however, our results suggest their relatively loose ΨL regulation in response to hydrologic stress occurs with a substantial hydraulic cost that may expose them to novel risks in a more drought-prone future. We end by discussing mechanisms that allow these species to tolerate and/or recover from hydraulic damage.

Recent results suggest that metabolism-mediated stomatal closure mechanisms are important to regulate differentially the stomatal speediness between ferns and angiosperms. However, evidence directly linking mesophyll metabolism and the slower stomatal conductance (gs) in ferns is missing. Here we investigated the effect of exogenous application of abscisic acid (ABA), sucrose and mannitol on gs kinetics and carried out a metabolic fingerprinting analysis of ferns and angiosperms leaves harvested throughout a diel course. Ferns stomata did not respond to ABA in the time period analysed. No differences in the relative decrease in gs was observed between ferns and the angiosperm following provision of sucrose or mannitol. However, ferns have slower gs responses to these compounds than angiosperms. Metabolomics analysis highlights that ferns have higher accumulation of secondary rather than primary metabolites throughout the diel course, with the opposite being observed in angiosperms. Our results indicate that metabolism-mediated stomatal closure mechanism is conserved among ferns and angiosperms and that the slower stomatal closure in ferns is associated to a reduced capacity to respond to mesophyll-derived sucrose and to a higher carbon allocation toward secondary metabolism, which likely modulates both photosynthesis-stomatal movements and growth-stress tolerance trade-offs.

Similar to other cropping systems, few walnut cultivars are used as scion in commercial production. Germplasm collections can be used to diversify cultivar options and hold potential for improving crop productivity, disease resistance and stress tolerance. In this study we explored the anatomical and biochemical bases of photosynthetic capacity in 11 J. regia accessions in the USDA-ARS National Clonal Germplasm Repository. Net assimilation rate (An) differed significantly among accessions and was greater in those from lower latitudes coincident with increases in stomatal and mesophyll conductance, leaf thickness, mesophyll porosity and gas-phase diffusion, and leaf nitrogen, and lower leaf mass and stomatal density. High CO2-saturated assimilation rates led to increases in An under limiting conditions. Greater An was found in lower latitude accessions native to climates with more frost-free days, greater precipitation seasonality, and lower temperature seasonality. As expected, water stress consistently impaired photosynthesis with the highest % reductions in three lower latitude accessions (A3, A5, and A9), which had the highest An under well-watered conditions. However, An for A3 and A5 remained amongst the highest under dehydration. J. regia accessions, which have leaf structural traits and biochemistry that enhance photosynthesis, could be used as commercial scions or breeding parents to enhance productivity.

Cold acclimation in plants is a complex phenomenon involving numerous stress-responsive transcriptional and metabolic pathways. Existing gene expression studies have primarily addressed cold acclimation responses in herbaceous plants, and few have focused on perennial evergreens, such as conifers, that survive extremely low temperatures during winter. Relative to Arabidopsis leaves, the main transcriptional response of Norway spruce (Picea abies (L.) H. Karst) needles exposed to cold was delayed, and this delay was associated with slower development of freezing tolerance. Despite this difference in timing, our results indicate that, similar to herbaceous species, Norway spruce principally utilizes early response transcription factors (TFs) of the APETALA 2/ethylene-responsive element binding factor (AP2/ERF) superfamily and NAM (no apical meristem)/ATAF (Arabidopsis Transcription Factors)/CUC (cup shaped cotyledon) (NACs). The needles and root of Norway spruce showed contrasting results, in keeping with their different metabolic and developmental states. Regulatory network analysis identified conserved TFs, including a root-specific bHLH101 homolog, and other members of the same TF family with a pervasive role in cold regulation, such as homologs of ICE1 and AKS3, and also homologs of the NAC (anac47 and anac28) and AP2/ERF superfamilies (DREB2 and ERF3), providing new functional insights into cold stress response strategies in Norway spruce.

To explore diversity in cold hardiness mechanisms, high resolution magnetic resonance imaging (MRI) was used to visualize freezing behaviors in wintering flower buds of Daphne kamtschatica var. jezoensis, which have no bud scales surrounding well-developed florets. MRI images showed that anthers remained stably supercooled to -14 ∼ -21°C or lower whilst most other tissues froze by -7°C. Freezing of some anthers detected in MRI images at ∼ -21°C corresponded with numerous low temperature exotherms and also with the “all-or-nothing” type of anther injuries. In ovules/pistils, only embryo sacs remained supercooled at -7°C or lower, but slowly dehydrated during further cooling. Cryomicroscopic observation revealed ice formation in the cavities of calyx tubes and pistils but detected no ice in embryo sacs or in anthers. The distribution of ice nucleation activity in floral tissues corroborated the tissue freezing behaviors. Filaments likely work as the ice blocking barrier that prevents ice intrusion from extracellularly frozen calyx tubes to connecting unfrozen anthers. Unique freezing behaviors were demonstrated in Daphne flower buds: preferential freezing avoidance in male and female gametophytes and their surrounding tissues (by stable supercooling in anthers and by supercooling with slow dehydration in embryo sacs) whilst the remaining tissues tolerate extracellular freezing.

Xylem embolism resistance varies across species influencing drought tolerance, yet little is known about the determinants of the embolism resistance of an individual conduit. Here we conducted an experiment using the optical vulnerability method to test whether individual conduits have a specific water potential threshold for embolism formation and whether pre-existing embolism in neighbouring conduits alters this threshold. Observations were made on a diverse sample of angiosperm and conifer species through a cycle of dehydration, rehydration and subsequent dehydration to death. Upon rehydration after the formation of embolism, no refilling was observed. When little pre-existing embolism was present, xylem conduits had a conserved, individual, embolism resistance threshold that varied across the population of conduits. The consequence of a variable conduit-specific embolism threshold is that a small degree of pre-existing embolism in the xylem results in an apparently more resistant xylem in a subsequent dehydration, particularly in angiosperms with vessels. While our results suggest that pit membranes separating xylem conduits are critical for maintaining a conserved individual embolism threshold for given conduit when little pre-exisiting embolism is present, as the percentage of embolized conduits increases, gas movement, local pressure differences, and connectivity between conduits increasingly contribute to embolism spread.

Xylem is a main road in plant long-distance communication. Through xylem plants transport water, minerals and myriad of signaling molecules. With the onset during early embryogenesis, the development of xylem tissues relays on hormone gradients, activity of unique transcription factors, distribution of mobile miRNAs and receptor-ligand pathways. These regulatory mechanisms are often interconnected and all together contribute to the plasticity of water conducting tissue. Remarkably, root xylem carries water to all above-ground organs and therefore influences all aspects of plant growth. Because of the global warming and increasing water deficit, we need to come up with solutions for the crops of the future. It is clear that structure of water conducting elements directly impacts water transport within the plant. Among plant pathogens- vascular wilts attacking xylem -are the most harmful. Our knowledge about xylem anatomy and rewiring ability could bring the solutions against these diseases. In this review we summarize the recent findings on the molecular mechanisms of xylem formation with a special attention to the cellular changes, and cell wall rearrangements that are necessary to create functional capillaries. We emphasize the impact of abiotic factors and pathogens on xylem plasticity and discuss multidisciplinary approach to model xylem in crops.