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.

Breeding drought stress tolerance is an integral part of our current and future goals of sustainable agricultural production. In the present study, we examined the natural variation of HvP5cs1 and demonstrated the utility of a wild barley allele for drought stress adaptation in cultivated barley. Sequencing the 5-end regulatory region among 49 barley accessions identified a genetically distinct allele of HvP5cs1 promoter from a wild barley ISR42-8. Allele mining of HvP5cs1 indicated quantitative variation in proline accumulation which was associated with promoter polymorphisms across the cluster of abscisic acid-responsive elements (ABRE), ABRE-related coupling elements, and MYB binding motifs. A near-isogenic line (NIL-143) harboring the HvP5cs1 allele from the highest proline accumulating wild barley ISR42-8 was developed in cultivated barley Scarlett through marker-assisted backcrossing (BC6). NIL-143 preserved the genetic competence of ISR42-8 to accumulate proline in higher concentrations under drought conditions at seedling and reproductive stages. Under drought stress, NIL-143 maintained superior membrane integrity, reduced pigment damage, and sustained photosynthetic health compared to Scarlett. NIL-143 presented a remarkable improvement in drought stress recovery than Scarlett. Further, the introgression line exhibited improved yield attributes, especially superior grain weight compared to Scarlett under field drought conditions. In conclusion, the present data uncover the genetic regulation of HvP5cs1 mediated proline accumulation and elucidate its role in drought stress adaptation and yield stability in barley.

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.

The isohydric-anisohydric continuum describes the relative stringency of stomatal control of leaf water potential ( ψleaf) during drought. Hydroscape area ( HA) – the water potential landscape over which stomata regulate ψleaf – has emerged as a useful metric of the iso/anisohydric continuum because it is strongly linked to several hydraulic, photosynthetic, and structural traits. Previous research on HA focused on broad ecological patterns involving several plant clades. Here we investigate relationships of HA to climatic conditions and functional traits across ecologically diverse but closely related species while accounting for phylogeny. Across a macroclimatic moisture gradient, defined by the ratio of mean annual precipitation to mean annual pan evaporation ( P/Ep), HA decreased with P/Ep for ten Eucalyptus species. Greater anisohydry reflects lower turgor loss points and greater hydraulic safety, mirroring global patterns. More isohydric species have mesophyll photosynthetic capacity that is more sensitive to ψleaf, consistent with an earlier model for optimal stomatal behavior. Hydroscapes exhibit little plasticity in response to variation in water supply, and the extent of plasticity does not vary with P/Ep of native habitats. These findings strengthen the case that HA is a useful metric for characterizing drought tolerance and water-status regulation.

Heat stress (HS) caused by ambient high temperature pose a threat to plants. In the natural and agricultural environment, plants often encounter repeated and changeable HS. Moderate HS primes plants to establishment of a molecular ‘thermomemory’ that enables plants to withstand a later-and possibly more extreme-HS attack. Recent years, brassinosteroids (BRs) have been implicated in HS response whereas little is known about whether BRs signal transduction modulates thermomemory. Here, we uncover the positive role of BRs signaling in thermomemory of Arabidopsis thaliana. Heat priming induces de novo synthesis and nuclear accumulation of BRI1-EMS-SUPPRESSOR (BES1), the key regulator of BRs signaling. BRs promote the accumulation of dephosphorylated BES1 during memory phase, blocking BRs synthesis impairs dephosphorylation. During HS memory, BES1 is required to maintain sustained induction of HS memory genes and directly targets APX2 and HSFA3 for activation. In summary, our results reveal a BES1-required, BRs-enhanced transcriptional control module of thermomemory in Arabidopsis thaliana.

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.

Iron (Fe) deficiency restricts crop yields in calcareous soil. Thus, a novel Fe chelator, proline-2′-deoxymugineic acid (PDMA), based on the natural phytosiderophore 2′-deoxymugineic acid (DMA), was developed to solve the Fe deficiency problem. However, the effects and mechanisms of PDMA relevant to the Fe nutrition and yield of dicots grown under field conditions require further exploration. In this study, pot and field experiments with calcareous soil were conducted to investigate the effects of PDMA on the Fe nutrition and yield of peanuts. The results demonstrate that PDMA could dissolve insoluble Fe in the rhizosphere and up-regulate expression of the yellow stripe-like family gene AhYSL1 to improve the Fe nutrition of peanuts. Moreover, the chlorosis and growth inhibition induced by Fe deficiency were significantly diminished. Importantly, under field conditions, the peanut yield and kernel micronutrition were notably promoted by PDMA application. Our results indicate that PDMA promotes the dissolution of insoluble Fe and a rich supply of Fe in the rhizosphere, increasing yields through integrated improvements in soil–plant Fe nutrition at the molecular and ecological levels. In conclusion, the efficacy of PDMA for improving the Fe nutrition and yield of peanut indicates its outstanding potential for agricultural applications.

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 regulation of protein synthesis plays an important role in growth and development in all organisms. Upstream open reading frames (uORFs) are commonly found in eukaryotic mRNA transcripts and typically attenuate the translation of associated downstream main ORFs (mORFs). Conserved peptide uORFs (CPuORFs) are a rare subset of uORFs, some of which have been shown to conditionally regulate translation by ribosome stalling. Here we identify three Arabidopsis CPuORFs of ancient origin that regulate translation of any downstream ORF, in response to agriculturally significant environmental signals: heat stress and water limitation. We provide evidence that different sequence classes of CPuORF stall ribosomes during different phases of translation and show that plant CPuORFs act as environmental sensors that can be utilised as inducible regulators of translation with broad application.

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.

Long non-coding RNAs (lncRNAs) have been considered to be important regulators of gene expression in a range of biological processes in plants. A large number of lncRNAs have been identified in plants. However, most of their biological functions still remain to be determined. Here, we identified total 3 004 lncRNAs in cassava under normal or cold-treated conditions from Iso-seq data. We further characterized a lincRNA, CRIR1, as a novel positive regulator of the plant response to cold stress. CRIR1 can be significantly induced by cold treatment. Overexpression of CRIR1 in cassava enhanced the cold tolerance of transgenic plants. Transcriptome analysis demonstrated that CRIR1 regulates a range of cold stress-related genes in a CBF-independent pathway. We further found that CRIR1 RNA can interact with MeCSP5, a homolog of the cold shock protein that acts as RNA chaperones, indicating that CRIR1 may recruit MeCSP5 to improve the translation efficiency of mRNA. In summary, our study greatly extends the repertoire of lncRNAs in plants as well as its responding to cold stress. Moreover, it reveals a sophisticated mechanism by which CRIR1 regulates plant cold stress response by modulating the expression of stress-responsive genes and increasing the translational yield.

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.

Oxylipins are lipid-derived molecules that are ubiquitous in eukaryotes and whose functions in plant physiology have been widely reported. They appear to play a major role in plant immunity by orchestrating reactive oxygen species (ROS) and hormone-dependent signalling pathways. The present work focuses on the specific case of fatty acid hydroperoxides (HPOs). Although some studies report their potential use as exogenous biocontrol agents for plant protection, evaluation of their efficiency in planta is lacking and no information is available about their mechanism of action. In this work, the potential of 13(S)-hydroperoxyoctadeca-(9Z,11E)-dienoic acid (13-HPOD) and 13(S)-hydroperoxy-(9Z,11E,15Z)-octadecatrienoic acid (13-HPOT), as plant defence elicitors and the underlying mechanism of action are investigated. Arabidopsis thaliana leaf resistance to Botrytis cinerea was observed after root application with HPOs. They also activate early immunity-related defence responses, like ROS. As previous studies have demonstrated their ability to interact with plant plasma membranes (PPM), we have further investigated the effects of HPOs on biomimetic PPM structure using complementary biophysics tools. Results show that HPO insertion into PPM impacts its global structure without solubilizing it. Relationship between biological assays and biophysical analysis suggests that lipid amphiphilic elicitors that directly act on membrane lipids might trigger early plant defence events