Singlet Oxygen (SO) is among the most potent reactive oxygen species, and readily oxidizes proteins, lipids, and DNA. It can be generated at the plant surface by phototoxins in the epidermis, acting as a direct defense against pathogens and herbivores (including humans). SO can also accumulate within mitochondria, peroxisomes, cytosol, and the nucleus through multiple enzymatic and non-enzymatic processes. However, the primary location of SO in plants is in the chloroplast, where it results from transfer of light energy from PhotosystemII to triplet oxygen. SO accumulates in response to diverse stresses that perturb chloroplast metabolism, and while its short half-life precludes exiting the chloroplast, it participates in retrograde signaling through the EXECUTER1 sensor, generation of carotenoid metabolites, and possibly other unknown pathways. SO thereby reprograms nuclear gene expression and modulates hormone signaling and programmed cell death. While SO signaling has long been known to regulate plant responses to high-light stress, recent literature also suggests a role in plant interactions with insects, bacteria, and fungi. The goals of this review are to provide a brief overview of SO, summarize evidence for its involvement in biotic stress responses, and discuss future directions for the study of SO in signaling and defense.

Reactive oxygen species are important signaling molecules that influence many aspects of plant biology. One way in which ROS influence plant growth and development is by modifying intercellular trafficking through plasmodesmata (PD). Viruses have evolved to use plasmodesmata for their local cell-to-cell spread between plant cells, so it is therefore not surprising that they have found ways to modulate ROS and redox signaling to optimize plasmodesmata function for their benefit. This review examines how intracellular signaling via ROS and redox pathways regulate intercellular trafficking via PD during development and stress. The relationship between viruses and ROS-redox systems, and the strategies viruses employ to control PD function by interfering with ROS-redox in plants is also discussed.

Due to their stationery nature, plants are exposed to a diverse range of biotic and abiotic stresses, of which heavy metals stress poses as one of the most detrimental abiotic stresses, targeting crucial and vital processes. Heavy metals instigate the over-production of reactive oxygen species (ROS), and in order to mitigate the adverse effects of ROS, plants induce multiple defence mechanisms. Besides the negative implications of overproduction of ROS, these molecules play a multitude of signaling roles in plants, acting as a central player in the complex signaling network of cells. One of the signaling mechanisms it is involved in is the mitogen-activated protein kinase (MAPK) cascade, a signaling pathway used to transduce extracellular stimuli into intracellular responses. Plant MAPKs have been implicated in signaling of stresses, phytohormones and cell cycle cues. However, the influence of various heavy metals on MAPKs activation has not been well documented. In this review, we will attempt to address and summarize several aspects related to various heavy metal-induced ROS signaling, how these signals activate the MAPK cascade and the downstream transcription factors that instigates the plants response to these heavy metals. Moreover, we will highlight a modern research methodology that could characterize the novel genes associated with MAPKs and their roles in heavy metal stress.

Stomata play a pivotal role in regulating gas exchange between terrestrial plants and the atmosphere controlling water and carbon cycles at organismal, ecosystem and global levels. Accordingly, our objective was to investigate the impact of ultraviolet-B radiation, a neglected environmental factor varying with ongoing global change, on stomatal morphology and function by means of a comprehensive meta-analysis. We found 45 peer-reviewed publications containing altogether 143 case studies for analysis. The overall UV effect at the leaf level is to decrease stomatal conductance, stomatal aperture and stomatal size, although stomatal density was increased. The significant decline in conductance is marked in short-term experiments, with more modest decreases noted in long-term UV studies. We found that short-term experiments in growth chambers are not representative of long-term field UV effects on stomatal conductance. Further, we found a stronger UV effect in grasses than in herbs, while the reduction of stomatal conductance was insignificant in trees. It is hypothesised that these alterations in stomatal function have important potential consequences for plant life. In the short term, UV-mediated stomatal closure may reduce transpiration and alleviate drought stress. However, in the long term more complex changes in stomatal aperture, size and density may reduce carbon sink capacity, and enhance leaf and surface warming, potentially exacerbating the negative effects of drought and/or heatwaves on plant ecosystems and endangering long-term plant survival.

Summary: The hindrance of kernel development caused by waterlogging stress (WS) is the direct reason for the peanut yield reduction. Currently, the mechanism of kernel filling responding to WS remains unknown. The waterlogging-sensitive variety Huayu 39 was subjected to WS for 3 days after 7 days after gynophores touched the ground (DAG), and the key stage of WS affecting kernel filling is 14, 21, and 28 DAG. WS decreased the average filling rate and kernel dry weight. Therefore, transcriptome sequencing and widely-targeted metabolomic analysis were conducted on kernel to elucidate the mechanism for the decrease in average filling rate under WS, revealing that overexpression of the gene encoding tryptophan decarboxylase ( AhTDC), which caused the accumulation of melatonin, reduced kernel weight. The sucrose transformation rate was limited by the crosstalk between melatonin and ethylene, thereby reducing the kernel filling rate and hindering kernel development. Our results are crucial for formulating measures to alleviate the negative impact of WS on peanut yield and quality, providing a basis for exploring high-yield and high-quality cultivation, molecular-assisted breeding, and waterlogging prevention.

With continued global warming, plants are forecast to increasingly experience abiotic stress(es). Stomata on leaf surfaces are the gatekeepers to plant interiors, regulating gaseous exchanges that are crucial for both photosynthesis and outward water release. To optimise future productivity, accurate modelling of how stomata govern plant-environment interactions will be crucial. Here, we synergise optical and thermal imaging data to enhance transpiration modelling during water and/or nitrogen stress. By utilising hyperspectral data and partial least squares regression analysis of six plant traits and fluxes in wheat ( Triticum aestivum), we have developed a new spectral vegetation index; the combined nitrogen and drought index (CNDI), which can be used to detect both water stress and/or nitrogen deficiency. Upon full stomatal closure during drought, CNDI reduces as leaf biochemistry changes unfold, and during a combined stress experiment (drought and nitrogen deficiency), this is reflected in CNDI showing a strong relationship with leaf relative water content ( r2 = 0.70). By incorporating CNDI transformed with a sigmoid function into thermal-based transpiration modelling, we have increased the accuracy of modelling water fluxes during abiotic stress. If employed using future remote sensing technologies, our findings have the potential to markedly improve agricultural water usage and yields.

Xylem conduits have lignified walls to resist crushing pressures. The thicker the double-wall ( T) relative to its maximum diameter ( D), the greater the collapse/implosion resistance. Having xylem that is more resistant than necessary incurs high costs and reduced flow, while having xylem not resistant enough may lead to catastrophic collapse under drought. Despite the importance of xylem implosion safety in determining plant drought resistance, it is still unclear how leaves scale Tx D to trade-off among implosion safety, flow efficiency, mechanical support, and construction cost. We measured T and D in over 7,000 leaf xylem conduits of 122 ferns and angiosperms species to investigate how the Tx D scaling varies across species, clades, habitats, growth forms, and vein orders. Overall, leaf xylem conduits grow wider than thicker, potentially resulting in high flow efficiency and lower cost, but at the expense of high vulnerability to implosion. Conduits seem particularly vulnerable to implosion in monocots, aquatic species and in species from hydric habitats, as well as in major veins. The absence of strong trade-offs within the leaf functional traits examined suggests that implosion safety at the whole-leaf level cannot be easily predicted by the sum of the individual conduits’ resistance to collapse.

In plants, salicylic acid (SA) hydroxylation regulates SA homoeostasis, playing an essential role during plant development and response to pathogens. This reaction is catalyzed by SA hydroxylase enzymes, which hydroxylate SA producing 2,3- dihydroxybenzoic acid (2,3-DHBA) and/or 2,5-dihydroxybenzoic acid (2,5-DHBA). Several SA hydroxylases have been recently identified and characterized from different plant species; however, no such activity has been previously reported in maize. In this work, we describe the identification and characterization of a new SA hydroxylase in maize plants. This enzyme, with high sequence similarity to previously described SA hydroxylases from Arabidopsis and rice, converts SA into 2,5-DHBA; however, it shows different kinetics properties to those from previously characterized enzymes, and it also catalyzes the conversion of the flavonoid dihydroquercetin into quercetin in in vitro activity assays, suggesting that the maize enzyme may have different roles in vivo as those previously reported from other species. Despite this, ZmS5H can complement the resistance to pathogen and early senescence phenotypes of Arabidopsis s3h mutant plants. Finally, we characterized a maize mutant in the S5H gene ( s5hMu) that has altered growth, senescence and increased resistance against Colletotrichum graminicola infection, showing not only changes in SA and 2,5-DHBA but also variations in flavonol levels. Together, the results presented here provide evidence that SA hydroxylases in different plant species have evolved to show differences in catalytic properties that may be important to fine tune SA levels and other phenolic compounds such as flavonols to regulate different aspects of plant development and defense against pathogens.

Day respiration ( R d) is the metabolic, non-photorespiratory process by which illuminated leaves liberate CO 2 during photosynthesis. R d is used routinely in photosynthetic models and is thus critical for calculations. However, metabolic details associated with R d are poorly known, and this can be problematic to predict how R d changes with environmental conditions and relates to night respiration. It is often assumed that day respiratory CO 2 release just reflects ‘ordinary’ catabolism (glycolysis and Krebs ‘cycle’). Here, we carried out a pulse-chase experiment, whereby a 13CO 2 pulse in the light was followed by a chase period in darkness and then in the light. We took advantage of non-targeted, isotope-assisted metabolomics to determine non-‘ordinary’ metabolism, detect carbon remobilisation, and compare light and dark 13C utilisation. We found that several concurrent metabolic pathways (‘ordinary’ catabolism, oxidative pentose phosphates pathway, amino acid production, nucleotide biosynthesis, and secondary metabolism) took place in the light and participate in net CO 2 efflux associated with day respiration. Flux reconstruction from metabolomics leads to an underestimation of R d, further suggesting the contribution of a variety of CO 2-evolving processes. Also, the cornerstone of the Krebs ‘cycle’, citrate, is synthetised de novo from photosynthates mostly in darkness, and remobilised or synthesised from stored material in the light. Collectively, our data provides direct evidence that leaf day respiration ( i) involves several CO 2-producing reactions and ( ii) is fed by different carbon sources, including stored carbon disconnected from current photosynthates.

The plant cell wall is a plastic structure of variable composition that constitutes the first line of defense against environmental challenges. Lodging and drought are two stressful conditions that severely impact on maize yield. In a previous work, we characterized the cell walls of two maize inbreds susceptible (EA2024) or resistant (B73) to stalk-lodging. Here, we show that drought induces phenotypical, physiological, cell wall, and transcriptional changes with distinct dynamics in the two inbreds, and that B73 is less tolerant than EA2024 to this stress. While in control conditions, stalk of EA2024 had higher levels of cellulose, uronic acids and p-coumarate than B73, upon drought these displayed increased levels of arabinose-enriched polymers, such as pectin-arabinans and arabinogalactan proteins, and a decreased lignin content. By contrast, a deeper rearrangement of cell walls including the modification of lignin composition and an increase of uronic acids was observed in B73. Drought induced higher changes in gene expression in B73 compared to EA2024, particularly in cell wall-related genes, that were altered in an inbred-specific manner. Transcription factor enrichment assays unveiled inbred-specific regulatory networks coordinating cell wall genes expression. Altogether, these findings reveal that B73 and EA2024 inbreds, with opposite stalk-lodging phenotypes, undertake different cell wall modification strategies in response to drought. We propose that the specific cell wall composition that confers lodging resistance to B73 compromises its cell wall plasticity and renders this inbred more susceptible to drought.

Nucleotide-binding, leucine-rich repeat (NLR) genes play a pivotal role in shaping plant effector-triggered immunity in response to pathogen invasions. However, the mechanisms governing the expression and behavior of NLRs, particularly in the context of head-to-head NLR gene pairs, in the presence of pathogens, remain uncovered. In this study, we dissected the Pik-H4 promoter (P Pik-H4) at the TATA boxes and conducted an in-depth investigation into split promoter activity using Agro-infiltration assays. The segments spanning 593-1232 bp and 2016-2492 bp (starting from -1 bp of Pik1-H4) within P Pik-H4 emerged as core regions for expressing Pik1-H4 and Pik1-H4 respectively. Nevertheless, merging these two core fragments failed to recover the promoter activity in both directions. Employing Gus staining, promoter activity assays and qRT-PCR, we unveiled the co-expression of Pik1-H4 and Pik2-H4 throughout the whole plant. Additionally, in the presence of the rice blast fungus, their co-amplification was observed in leaves and leaf sheaths. Strikingly, Pik-H4 exhibited heightened expression within vascular bundles. Moreover, perturbing the Pik1-H4 and Pik2-H4 co-expression relationship via overexpression in rice or Nicotiana did not disrupt the immune response. Upon infection, the singleton Pik 1-H4 localized within vesicles, while Pik 2-H4 predominantly occupied the nucleus within leaf sheath cells. Transcriptome analysis highlighted Pik-H4-mediated resistance triggering a transcriptome reprogramming between 12 and 24 hours post-inoculation. Notably, overexpression of Pik1-H4 or Pik2-H4 enriches various pathways compared to the Pik-H4 Lijiangheituanxingu near-isogenic lines. In summary, these findings unravel the intricate dynamics of co-expression and singular functionality within NLR bidirectional gene pairs upon pathogen invasion.

High temperature negatively impacts the yield and quality of fruit crops. Exogenous melatonin (MT) application has shown the capability to enhance heat tolerance, but the response of endogenous MT to heat stress, particularly in perennial fruit trees, remains elusive. This study investigated the effects of high temperatures on transgenic apple plants overexpressing the MT biosynthetic gene N-acetylserotonin methyltransferase 9 ( MdASMT9). Endogenous MT protected transgenic plants from heat stress, scavenging reactive oxygen species (ROS) and increasing soluble carbohydrates and amino acids levels. MdASMT9-overexpressing plants also maintained higher photosynthetic activity by protecting the chloroplasts from damage. Transcriptome sequencing indicates that MdASMT9 overexpression promoting the expression of HSFA1d, HSFA2-like, and HSFA9b, and inhibiting the transcription of HSFB1 and HSFB2b. Application of MT and overexpression of MdASMT9 reduced abscisic acid (ABA) accumulation through promoting MdWRKY33-mediated transcriptional inhibition of MdNCED1 and MdNCED3, thus promoting stomatal opening for better heat dissipation. Furthermore, melatonin enhanced autophagic activity through promoting MdWRKY33-mediated transcriptional enhancement of MdATG18a under heat stress . These findings provide new sight into the regulation of endogenous MT and its role in improving heat tolerance in perennial fruit trees.

Photosynthesis is the foundation of all life on Earth, providing oxygen and energy. However, if not well regulated, it can also generate toxic reactive oxygen species (ROS), which can cause photodamage. Regulation of photosynthesis is highly dynamic, responding to both environmental and metabolic cues, and occurs at many levels, from light capture to energy storage and metabolic processes. One general mechanism of regulation involves the reversible oxidation and reduction of protein thiol groups, which can affect the activity of enzymes and the stability of proteins. Such redox regulation has been well studied in stromal enzymes, but more recently evidence has emerged of redox control of thylakoid lumenal enzymes. This review/hypothesis paper summarizes the latest research and discusses several open questions and challenges to achieving effective redox control in the lumen, focusing on the distinct environments and regulatory components of the thylakoid lumen, including the need to transport electrons across the thylakoid membrane, the effects of pH changes in the stromal and lumenal compartments, and the observed differences in redox states. These constraints suggest that activated oxygen species are likely to be major regulatory contributors to lumenal thiol redox regulation, with key components and processes yet to be discovered.

Wood serves crucial functions in plants, yet our understanding of the molecular regulation governing the composition, arrangement, and dimensions of its cells remains limited. The abrupt change in wood anatomy of lianas represents an excellent model to address the underlying mechanism, although consistent triggering factors for this process remain uncertain. In this study we examined how physical support attachment impacts the development of lianescent xylem anatomy in Bignonia magnifica (Bignoniaceae), employing a comprehensive approach integrating detailed anatomical analysis with gene expression profiling of cambium and differentiating xylem. Our findings demonstrate that attachment to physical supports triggers the formation of lianescent xylem, leading to increased vessel size, range of vessel sizes, broader vessel distribution, reduced fiber content, and higher potential specific water conductivity. These shifts in wood anatomy coincide with the downregulation of genes associated with cell division and cell wall biosynthesis, and the upregulation of transcription factors (TFs), defense/cell death, and hormone-responsive genes in the lianescent xylem. Based on our results, we propose a model delineating the molecular control underlying the formation of lianescent xylem, revealing how the increased complexity of lianescent anatomy reflects a more intricate transcriptional regulatory network encompassing a more diverse repertoire of TFs and hormone-responsive genes.

Leaf gas exchange measurements provide an important tool for inferring a plant’s photosynthetic biochemistry. In most cases, the responses of photosynthetic CO 2 assimilation to variable intercellular CO 2 concentrations ( A/ Ci response curves) are used to model the maximum rate of carboxylation by ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco, V cmax) and the rate of electron transport at a given photosynthetically active radiation (PAR; J PAR). The standard Farquhar-Von Caemmerer-Berry model is typically used with default parameters of Rubisco kinetic values and mesophyll conductance to CO 2 ( g m) derived from tobacco that impairs analytical reliability across species. To study this, here we measured the temperature responses of key in vitro Rubisco catalytic properties and g m in cotton ( Gossypium hirsutum cv. Sicot 71) and derived V cmax and J 2000 ( J at 2000 µmol m -2 s -1 PAR) from cotton A/ Ci curves incrementally measured at 15°C to 40°C using cotton and tobacco parameters with our new automated fitting R package ‘OptiFitACi’. When applied to cotton, the tobacco parameters produced unrealistic J 2000: V cmax ratio of <1 at 25°C, two- to three-fold higher estimates of V cmax, approximately 50% higher estimates of J 2000 and more variable estimates of V cmax and J 2000, compared to model parameterisation with cotton-derived values. We determined that errors arise when using a g m of 0.23 mol m -2 s -1 bar -1 or below and Rubisco CO 2-affinities under ambient O 2 ( K C 21%O2) outside 461 µbar to 627 µbar to model A/ C i responses in cotton. We show how the multi- A/ C i modelling capabilities of ‘OptiFitACi’ serves as a robust, user-friendly extension of ‘plantecophys’ by providing simplified temperature-sensitivity and species-specificity parameterisation capabilities to enable higher accuracy estimates of V cmax and J 2000.

RNA editing is a tightly controlled process by which cytidines are converted to uridines in RNAs transcribed from the chloroplast and mitochondrial genomes in flowering plants. Multiple organellar RNA editing factor (MORF) complex was recently shown to be highly associated with C-to-U RNA editing activity of vascular plant editosome. However, mechanisms by which MORF9 mediates plastid RNA editing to control plant development in response to environmental cues remains obscure. In this study, we found that loss of MORF9 function impaired PSII efficiency, NDH activity, and carbohydrate production, rapidly promoted nuclear gene expression including sucrose transporter and sugar/energy responsive genes, and attenuated seedling development under sugar starvation conditions. Sugar repletion increased MORF9 and MORF2 expression in wild-type seedlings and promoted inefficiency of matK-706C, accD-794C, ndhD-383C and ndhF-290C RNA editing in morf9 mutant. This RNA editing inefficiency was associated with altered cell division in root meristem zone and nuclear gene expression in the morf9 mutant. Using gin2, snrk1, morf9 single and double mutants and overexpression of SnRK1 (KIN10) or HXK1 in the morf9 mutant background demonstrated that RNA editing efficiency of ndhD-383C and ndhF-290C sites was diminished in the gin2/morf9 double mutants, and editing efficiency of matK-706C, accD-794C, ndhD-383C and ndhF-290C sites was significantly diminished in the snrk1/morf9 double mutants. Overexpressing HXK1 or SnRK1 promoted RNA editing rate of matK, accD, ndhD, and ndhF in leaves of morf9 mutants，indicating that HXK1 might be required for MORF9 mediated ndhD-383C and ndhF-290C editing, while SnRK1 may only be required for MORF9 mediated ndhF-290C site editing. Collectively these findings suggest that sugar and/or its intermediary metabolites impair MORF9 mediated plastid RNA editing resulting in derangements of plant root development.

D-amino acids are the D stereoisomers of common L-amino acids found in proteins. In the past two decades, the occurrence of D-amino acids in plants has been reported and circumstantial evidence for a role in several processes has been provided, including the interaction with soil microorganisms or an interference with cellular signalling. However, examples are relatively scarce and D-amino acids can also be detrimental, some of them inhibiting growth and development. Thus, the persistence of a D-amino acid metabolism in plants is rather surprising and evolutive origins of D-amino acid metabolism is presently unclear. Systemic analysis of sequences associated with enzymes of D-amino acid metabolism shows that they are not simply inherited from cyanobacterial metabolism. In effect, the history of enzymes of plant D-amino acid metabolism likely involves several steps, cellular compartments, gene transfers and losses. Regardless of evolutive steps, enzymes of D-amino acid metabolism like D-amino acid transferases or racemases have been kept by higher plants and not simply eliminated, hence it is likely that they fulfil important metabolic roles, which can be illustrated with serine, tryptophan, and folate metabolism. We suggest that D-amino acid metabolism was perhaps crucial to support metabolic functions required during land plants evolution.

Early flowering is a survival strategy in wheat ( Triticum aestivum L.) that sacrifices grain yield under long photoperiod conditions, and this contradiction is greatly affected by floral growth and development. However, little is known about the regulatory mechanisms that remove the barrier between “early flowering” and “high yielding” during floret development. Here, we showed high-resolution analyses of the number and morphology of floret primordia and the transcriptomes of wheat spikes in three light regimens. The development of all floret primordia in a spike could be divided into four distinct stages: differentiation (Stage I), differentiation and morphology development concurrently (Stage II), morphology development (Stage III), and polarization (Stage IV). Compared to the controls, the long photoperiod supplemented with red light treatment shortened the time required to complete Stage I-II, then improved assimilates in the spike and promoted anther development, thereby increasing fertile floret primordia during Stage III, and maintained fertile floret primordia development during Stage IV until they became fertile florets (grains) via a dynamic gene network centered on ubiquitin, calcium signaling, aldehyde dehydrogenase, zinc finger proteins, and heat shock proteins. Our findings proposed a light regimen, critical stages, and candidate regulators that promoted early flowering and high yield in wheat.