Introduction of a terminal electron sink in chloroplasts decreases leaf cell expansion associated with higher proteasome activity and lower endoreduplication.

Foliar development involves successive phases of cell proliferation and expansion that determine the final leaf size, and is characterized by an early burst of reactive oxygen species generated in the photosynthetic electron transport chain (PETC). Introduction of the alternative PETC acceptor flavodoxin in tobacco chloroplasts led to a reduction in leaf size associated to lower cell expansion, without affecting cell numbers per leaf. Proteomic analysis showed that components of the light-harvesting systems accumulated before electron-transport proteins, suggesting a mechanism for the early oxidative event. Flavodoxin expression did not affect biogenesis of the PETC but prevented hydroperoxide build-up through its function as electron sink. Mature leaves from flavodoxin-expressing plants were shown to contain higher levels of transcripts encoding components of the proteasome, a key negative modulator of organ size. Proteome profiling revealed that this differential accumulation initiated during expansion and led to increased proteasomal activity, whereas a proteasome inhibitor reverted the flavodoxin-dependent size phenotype. Cells expressing plastid-targeted flavodoxin displayed lower endoreduplication, also associated to decreased organ size. These results provide novel insights into the regulation of leaf growth by chloroplast-generated redox signals, and highlight the potential of alternative electron shuttles to investigate the link(s) between photosynthesis and plant development.

Transcriptomic and metabolomic approaches elucidate the systemic response of wheat plants under waterlogging.

Extreme weather conditions lead to significant imbalances in crop productivity, which in turn affect food security. Flooding events cause serious problems for many crop species such as wheat. Although metabolic readjustments under flooding are important for plant regeneration, underlying processes remain poorly understood. Here, we investigated the systemic response of wheat to waterlogging using metabolomics and transcriptomics. A 12 d exposure to excess water triggered nutritional imbalances and disruption of metabolite synthesis and translocation, reflected by reductions in plant biomass and growth performance. Metabolic and transcriptomic profiling in roots, xylem sap, and leaves indicated anaerobic fermentation processes as a local response in roots. Differentially expressed genes and ontological categories revealed that carbohydrate metabolism plays an important role in the systemic response. Analysis of the composition of xylem exudates revealed decreased root-to-shoot translocation of nutrients, hormones, and amino acids. Interestingly, among all metabolites measured in xylem exudates, alanine was the most abundant. Immersion of excised leaves derived from waterlogged plants in alanine solution led to increased leaf glucose concentration. Our results suggest an important role of alanine not only as an amino-nitrogen donor but also as a vehicle for carbon skeletons to produce glucose de novo and meet the energy demand during waterlogging.

Enhancing Maize Productivity and Soil Health under Salt Stress through Physiological Adaptation and Metabolic Regulation Using Indigenous Biostimulants.

Salinity poses a persistent threat to agricultural land, continuously jeopardizing global food security. This study aimed to enhance sweet corn (SC) fitness under varying levels of salinity using indigenous biostimulants (BioS) and to assess their impacts on plant performance and soil quality. The experiment included control (0 mM NaCl), moderate stress (MS; 50 mM NaCl), and severe stress (SS; 100 mM NaCl) conditions. Indigenous biostimulants, including compost (C), Bacillus sp., Bacillus subtilis (R), and a consortium of arbuscular mycorrhizal fungi (A) were applied either individually or in combination. Growth traits, physiological and biochemical parameters in maize plants, and the physico-chemical properties of their associated soils were assessed. SS negatively affected plant growth and soil quality. The RC combination significantly improved plant growth under SS, increasing aerial (238%) and root (220%) dry weights compared to controls. This treatment reduced hydrogen peroxide by 54% and increased peroxidase activity by 46% compared to controls. The indigenous biostimulants, particularly C and R, enhanced soil structure and mineral composition (K and Mg). Soil organic carbon and available phosphorus increased notably in C-treated soils. Furthermore, RC (437%) and CAR (354%) treatments exhibited a significant increase in glomalin content under SS. Indigenous biostimulants offer a promising strategy to mitigate salinity-related threats to agricultural land. They improve plant fitness, fine-tune metabolism, and reduce oxidative stress. In addition, the biostimulants improved the soil structure and mineral composition, highlighting their potential for reconstitution and sustainability in salt-affected areas. This approach holds promise for addressing salinity-related threats to global food security.

A non-canonical function of Arabidopsis ERECTA proteins and a role of the SWI3B subunit of the SWI/SNF chromatin remodeling complex in gibberellin signaling.

The Arabidopsis ERECTA family (ERf) of leucine-rich repeat receptor-like kinases (LRR-RLKs) comprising ERECTA (ER), ERECTA-LIKE 1 (ERL1), and ERECTA-LIKE 2 (ERL2) controls epidermal patterning, inflorescence architecture, and stomata development and patterning. These proteins are reported to be plasma membrane associated. Here we show that the er/erl1/erl2 mutant exhibits impaired gibberellin (GA) biosynthesis and perception alongside broad transcriptional changes. The ERf kinase domains were found to localize to the nucleus where they interact with the SWI3B subunit of the SWI/SNF chromatin remodeling complex (CRCs). The er/erl1/erl2 mutant exhibits reduced SWI3B protein level and affected nucleosomal chromatin structure. Similar to swi3c and brm plants with inactivated subunits of SWI/SNF CRCs, it also does not accumulate DELLA RGA and GAI proteins. The ER kinase phosphorylates SWI3B in vitro, and the inactivation of all ERf proteins leads to the decreased phosphorylation of SWI3B protein in vivo. The identified correlation between DELLA overaccumulation and SWI3B proteasomal degradation, and the physical interaction of SWI3B with DELLA proteins indicate an important role of SWI3B-containing SWI/SNF CRCs in gibberellin signaling. Co-localization of ER and SWI3B on GID1 (GIBBERELLIN INSENSITIVE DWARF 1) DELLA target gene promoter regions and abolished SWI3B binding to GID1 promoters in er/erl1/erl2 plants supports the conclusion that ERf-SWI/SNF CRC interaction is important for transcriptional control of GA receptors. Thus, the involvement of ERf proteins in the transcriptional control of gene expression, and observed similar features for human HER2 (epidermal growth family receptor member), indicate an exciting target for further studies of evolutionarily conserved non-canonical functions of eukaryotic membrane receptors.

A molecular framework for grain number determination in barley.

Flowering plants with indeterminate inflorescences often produce more floral structures than they require. We found that floral primordia initiations in barley (Hordeum vulgare L.) are molecularly decoupled from their maturation into grains. While initiation is dominated by flowering-time genes, floral growth is specified by light signaling, chloroplast, and vascular developmental programs orchestrated by barley CCT MOTIF FAMILY 4 (HvCMF4), which is expressed in the inflorescence vasculature. Consequently, mutations in HvCMF4 increase primordia death and pollination failure, mainly through reducing rachis greening and limiting plastidial energy supply to developing heterotrophic floral tissues. We propose that HvCMF4 is a sensory factor for light that acts in connection with the vascular-localized circadian clock to coordinate floral initiation and survival. Notably, stacking beneficial alleles for both primordia number and survival provides positive implications on grain production. Our findings provide insights into the molecular underpinnings of grain number determination in cereal crops.

A transaminase with ß-activity from Variovorax boronicumulans for the production of enantiopure ß-amino acids.

Enantioselective transamination of amino acids is a great challenge in biotechnology as suitable enzymes with wide substrate spectrum are rare. Here, we present a new transaminase from Variovorax boronicumulans (VboTA, Variovorax boronicumulansω-transaminase) which is specific for ß-amino acids. The amino acid sequence of VboTA is similar to an ω-transaminase from Variovorax paradoxus, for which a crystal-structure is available. This similarity is allowing us to classify VboTA as a fold type 1 ω-transaminase (ω-TA). Although both enzymes have a high sequence similarity (86% identities, 92% positives), there are differences in the active center, which allow VboTA to accept a broader substrate spectrum. Both enzymes have also a different temperature stability and temperature optimum. VboTA deaminates the D-form of aromatic ß-amino acids, such as ß-homophenylalanine and ß-phenylalanine as well as aliphatic ß-amino acids, such as ß-homoalanine and ß-leucine. The optimal reaction conditions turned out to be 32 °C and pH 9. Kinetic resolution lead to high enantiomeric excess of 86.6% to >99.9%, depending on the amino donor/acceptor pair. In contrast to many other ω-TAs, VboTA has a broad substrate spectrum and uses both aromatic or aliphatic amino acids. With γ-amino acids as substrates, VboTA showed no activity at all.

Arbuscular mycorrhizal colonization leads to a change of hormone profile in micropropagated plantlet Satureja khuzistanica Jam.

Phytohormones are supposed to contribute to the establishment of mutualistic Arbuscular mycorrhiza (AM) symbioses. However, their role in the acclimation of micropropagated plantlet inoculated with AM is still unknown. To address this question, we performed a hormone profiling during the acclimation of Satureja khuzistanica plantlets inoculated with Rhizoglomus fasciculatum. The levels of indoleacetic acid (IAA), methyl indole acetic acid, cis-zeatin, cis zeatin ribose, jasmonate, jasmonoyl isoleucine, salicylic acid, abscisic acid (ABA) were analyzed. Further, the relative gene expression of AOS (Allene oxide synthase) as a key enzyme of jasmonate biosynthesis, in either inoculated or non-inoculated micropropagated plantlets was evaluated during acclimation period. The concentrations of IAA and cis-zeatin increased in the plantlets inoculated by AM whereas the concentration of ABA decreased upon 60 days acclimation in the whole shoot of plantlets of S. khuzistanica. The relative expression of AOS gene resulted in an increase of isoleucine jasmonate, the bioactive form of jasmonate. Based on our results, IAA and cis-zeatin probably contribute to maintaining growth, and AM reduces transition stress by modifying ABA and jasmonate concentrations.

Comparative Transcriptome Analysis Unveils the Molecular Mechanism Underlying Sepal Colour Changes under Acidic pH Substratum in Hydrangea macrophylla

The hydrangea (Hydrangea macrophylla (Thunb). Ser.), an ornamental plant, has good marketing potential and is known for its capacity to change the colour of its inflorescence depending on the pH of the cultivation media. The molecular mechanisms causing these changes are still uncertain. In the present study, transcriptome and targeted metabolic profiling were used to identify molecular changes in the RNAome of hydrangea plants cultured at two different pH levels. De novo assembly yielded 186,477 unigenes. Transcriptomic datasets provided a comprehensive and systemic overview of the dynamic networks of the gene expression underlying flower colour formation in hydrangeas. Weighted analyses of gene co-expression network identified candidate genes and hub genes from the modules linked closely to the hyper accumulation of Al3+ during different stages of flower development. F3'5'H, ANS, FLS, CHS, UA3GT, CHI, DFR, and F3H were enhanced significantly in the modules. In addition, MYB, bHLH, PAL6, PAL9, and WD40 were identified as hub genes. Thus, a hypothesis elucidating the colour change in the flowers of Al3+-treated plants was established. This study identified many potential key regulators of flower pigmentation, providing novel insights into the molecular networks in hydrangea flowers.

Response of the wheat mycobiota to flooding revealed substantial shifts towards plant pathogens.

Rainfall extremes are intensifying as a result of climate change, leading to increased flood risk. Flooding affects above- and belowground ecosystem processes, representing a substantial threat to crop productivity under climate change. Plant-associated fungi play important roles in plant performance, but their response to abnormal rain events is unresolved. Here, we established a glasshouse experiment to determine the effects of flooding stress on the spring wheat-mycobiota complex. Since plant phenology could be an important factor in the response to hydrological stress, flooding was induced only once and at different plant growth stages, such as tillering, booting and flowering. We assessed the wheat mycobiota response to flooding in three soil-plant compartments (phyllosphere, roots and rhizosphere) using metabarcoding. Key soil and plant traits were measured to correlate physiological plant and edaphic changes with shifts in mycobiota structure and functional guilds. Flooding reduced plant fitness, and caused dramatic shifts in mycobiota assembly across the entire plant. Notably, we observed a functional transition consisting of a decline in mutualist abundance and richness with a concomitant increase in plant pathogens. Indeed, fungal pathogens associated with important cereal diseases, such as Gibberella intricans, Mycosphaerella graminicola, Typhula incarnata and Olpidium brassicae significantly increased their abundance under flooding. Overall, our study demonstrate the detrimental effect of flooding on the wheat mycobiota complex, highlighting the urgent need to understand how climate change-associated abiotic stressors alter plant-microbe interactions in cereal crops.

Cytokinins as boron deficiency signals to sustain shoot development in boron-efficient oilseed rape.

Boron (B) deficiency is a highly prominent nutrient disorder. While B-efficient accessions have recently been identified in the highly B-demanding crop oilseed rape, it remained unclear which physiological processes underlie B efficiency and which signaling pathways trigger an efficient B-deficiency response. Here, we compared, under three different B supply conditions, two Brassica napus accessions with contrasting B efficiency. Shoot biomass formation, B distribution patterns and metabolic dynamics of different phytohormone species were studied using a combination of mass spectrometry-based analyses and physiological measurements. Our results show that the B-efficient accession CR2267 does not differ from the B-inefficient accession CR2262 in terms of B accumulation and subcellular B-partitioning, although it displays no morphological B-deficiency symptoms under severe B-deficient conditions. Investigating phytohormone metabolism revealed a strong accumulation of cytokinins in CR2267 at a developmental stage when striking B-dependent differences in biomass and organ formation emerge in the two B. napus accessions. In contrast, elevated levels of the stress hormone abscisic acid as well as bioactive auxins, representing functional antagonists of cytokinins in shoots, were detected only in CR2262. Our results indicate that superior B efficiency in CR2267 relies on a higher B utilization efficiency that builds on an earlier and higher cytokinin biosynthesis required for the maintenance of the shoot meristem activity and proper leaf development. We further conclude that an elevated abundance of cytokinins is not a consequence of better plant growth but rather a presumption for better plant growth under low-B conditions.

Leaf bacterial microbiota response to flooding is controlled by plant phenology in wheat (Triticum aestivum L.).

Leaf microbiota mediates foliar functional traits, influences plant fitness, and contributes to various ecosystem functions, including nutrient and water cycling. Plant phenology and harsh environmental conditions have been described as the main determinants of leaf microbiota assembly. How climate change may modulate the leaf microbiota is unresolved and thus, we have a limited understanding on how environmental stresses associated with climate change driven weather events affect composition and functions of the microbes inhabiting the phyllosphere. Thus, we conducted a pot experiment to determine the effects of flooding stress on the wheat leaf microbiota. Since plant phenology might be an important factor in the response to hydrological stress, flooding was induced at different plant growth stages (tillering, booting and flowering). Using a metabarcoding approach, we monitored the response of leaf bacteria to flooding, while key soil and plant traits were measured to correlate physiological plant and edaphic factor changes with shifts in the bacterial leaf microbiota assembly. In our study, plant growth stage represented the main driver in leaf microbiota composition, as early and late plants showed distinct bacterial communities. Overall, flooding had a differential effect on leaf microbiota dynamics depending at which developmental stage it was induced, as a more pronounced disruption in community assembly was observed in younger plants.

A Chimeric TGA Repressor Slows Down Fruit Maturation and Ripening in Tomato.

The bZIP transcription factor (TF) SlTGA2.2 was previously highlighted as a possible hub in a network regulating fruit growth and transition to ripening (maturation phase). It belongs to a clade of TFs well known for their involvement in the regulation of the salicylic acid-dependent systemic acquired resistance. To investigate if this TGA TF plays a role in tomato fruit growth and maturation, we took advantage of the fruit-specific SlPPC2 promoter (PPC2pro) to target the expression of a SlTGA2.2-SRDX chimeric repressor in a developmental window restricted to early fruit growth and maturation. Here, we show that this SlTGA2.2-SRDX repressor alters early fruit development and metabolism, including chloroplast number and structure, considerably extends the time necessary to reach the mature green stage and slows down fruit ripening. RNA sequencing and plant hormone analyses reveal that PPC2pro:SlTGA2.2-SRDX fruits are maintained in an immature stage as long as PPC2pro is active, through early modifications of plant hormonal signaling and down-regulation of MADS-RIN and NAC-NOR ripening regulators. Once PPC2pro becomes inactive and therefore SlTGA2.2-SRDX expression is reduced, ripening can proceed, albeit at a slower pace than normal. Altogether, this work emphasizes the developmental continuum between fruit growth, maturation and ripening and provides a useful tool to alter and study the molecular bases of tomato fruit transition to ripening.

Flooding Causes Dramatic Compositional Shifts and Depletion of Putative Beneficial Bacteria on the Spring Wheat Microbiota.

Flooding affects both above- and below-ground ecosystem processes, and it represents a substantial threat for crop and cereal productivity under climate change. Plant-associated microbiota play a crucial role in plant growth and fitness, but we still have a limited understanding of the response of the crop-microbiota complex under extreme weather events, such as flooding. Soil microbes are highly sensitive to abiotic disturbance, and shifts in microbial community composition, structure and functions are expected when soil conditions are altered due to flooding events (e.g., anoxia, pH alteration, changes in nutrient concentration). Here, we established a pot experiment to determine the effects of flooding stress on the spring wheat-microbiota complex. Since plant phenology could be an important factor in the response to hydrological stress, flooding was induced only once and at different plant growth stages (PGSs), such as tillering, booting and flowering. After each flooding event, we measured in the control and flooded pots several edaphic and plant properties and characterized the bacterial community associated to the rhizosphere and roots of wheat plant using a metabarcoding approach. In our study, flooding caused a significant reduction in plant development and we observed dramatic shifts in bacterial community composition at each PGS in which the hydrological stress was induced. However, a more pronounced disruption in community assembly was always shown in younger plants. Generally, flooding caused a (i) significant increase of bacterial taxa with anaerobic respiratory capabilities, such as members of Firmicutes and Desulfobacterota, (ii) a significant reduction in Actinobacteria and Proteobacteria, (iii) depletion of several putative plant-beneficial taxa, and (iv) increases of the abundance of potential detrimental bacteria. These significant differences in community composition between flooded and control samples were correlated with changes in soil conditions and plant properties caused by the hydrological stress, with pH and total N as the soil, and S, Na, Mn, and Ca concentrations as the root properties most influencing microbial assemblage in the wheat mircobiota under flooding stress. Collectively, our findings demonstrated the role of flooding on restructuring the spring wheat microbiota, and highlighted the detrimental effect of this hydrological stress on plant fitness and performance.

Engineering Climate-Change-Resilient Crops: New Tools and Approaches.

Environmental adversities, particularly drought and nutrient limitation, are among the major causes of crop losses worldwide. Due to the rapid increase of the world's population, there is an urgent need to combine knowledge of plant science with innovative applications in agriculture to protect plant growth and thus enhance crop yield. In recent decades, engineering strategies have been successfully developed with the aim to improve growth and stress tolerance in plants. Most strategies applied so far have relied on transgenic approaches and/or chemical treatments. However, to cope with rapid climate change and the need to secure sustainable agriculture and biomass production, innovative approaches need to be developed to effectively meet these challenges and demands. In this review, we summarize recent and advanced strategies that involve the use of plant-related cyanobacterial proteins, macro- and micronutrient management, nutrient-coated nanoparticles, and phytopathogenic organisms, all of which offer promise as protective resources to shield plants from climate challenges and to boost stress tolerance in crops.

ITPK1 is an InsP6/ADP phosphotransferase that controls phosphate signaling in Arabidopsis.

In plants, phosphate (Pi) homeostasis is regulated by the interaction of PHR transcription factors with stand-alone SPX proteins, which act as sensors for inositol pyrophosphates. In this study, we combined different methods to obtain a comprehensive picture of how inositol (pyro)phosphate metabolism is regulated by Pi and dependent on the inositol phosphate kinase ITPK1. We found that inositol pyrophosphates are more responsive to Pi than lower inositol phosphates, a response conserved across kingdoms. Using the capillary electrophoresis electrospray ionization mass spectrometry (CE-ESI-MS) we could separate different InsP7 isomers in Arabidopsis and rice, and identify 4/6-InsP7 and a PP-InsP4 isomer hitherto not reported in plants. We found that the inositol pyrophosphates 1/3-InsP7, 5-InsP7, and InsP8 increase several fold in shoots after Pi resupply and that tissue-specific accumulation of inositol pyrophosphates relies on ITPK1 activities and MRP5-dependent InsP6 compartmentalization. Notably, ITPK1 is critical for Pi-dependent 5-InsP7 and InsP8 synthesis in planta and its activity regulates Pi starvation responses in a PHR-dependent manner. Furthermore, we demonstrated that ITPK1-mediated conversion of InsP6 to 5-InsP7 requires high ATP concentrations and that Arabidopsis ITPK1 has an ADP phosphotransferase activity to dephosphorylate specifically 5-InsP7 under low ATP. Collectively, our study provides new insights into Pi-dependent changes in nutritional and energetic states with the synthesis of regulatory inositol pyrophosphates.

Expression of Flavodiiron Proteins Flv2-Flv4 in Chloroplasts of Arabidopsis and Tobacco Plants Provides Multiple Stress Tolerance.

With the notable exception of angiosperms, all phototrophs contain different sets of flavodiiron proteins that help to relieve the excess of excitation energy on the photosynthetic electron transport chain during adverse environmental conditions, presumably by reducing oxygen directly to water. Among them, the Flv2-Flv4 dimer is only found in ß-cyanobacteria and induced by high light, supporting a role in stress protection. The possibility of a similar protective function in plants was assayed by expressing Synechocystis Flv2-Flv4 in chloroplasts of tobacco and Arabidopsis. Flv-expressing plants exhibited increased tolerance toward high irradiation, salinity, oxidants, and drought. Stress tolerance was reflected by better growth, preservation of photosynthetic activity, and membrane integrity. Metabolic profiling under drought showed enhanced accumulation of soluble sugars and amino acids in transgenic Arabidopsis and a remarkable shift of sucrose into starch, in line with metabolic responses of drought-tolerant genotypes. Our results indicate that the Flv2-Flv4 complex retains its stress protection activities when expressed in chloroplasts of angiosperm species by acting as an additional electron sink. The flv2-flv4 genes constitute a novel biotechnological tool to generate plants with increased tolerance to agronomically relevant stress conditions that represent a significant productivity constraint.

Transcriptional and Metabolic Profiling of Potato Plants Expressing a Plastid-Targeted Electron Shuttle Reveal Modulation of Genes Associated to Drought Tolerance by Chloroplast Redox Poise.

Water limitation represents the main environmental constraint affecting crop yield worldwide. Photosynthesis is a primary drought target, resulting in over-reduction of the photosynthetic electron transport chain and increased production of reactive oxygen species in plastids. Manipulation of chloroplast electron distribution by introducing alternative electron transport sinks has been shown to increase plant tolerance to multiple environmental challenges including hydric stress, suggesting that a similar strategy could be used to improve drought tolerance in crops. We show herein that the expression of the cyanobacterial electron shuttle flavodoxin in potato chloroplasts protected photosynthetic activities even at a pre-symptomatic stage of drought. Transcriptional and metabolic profiling revealed an attenuated response to the adverse condition in flavodoxin-expressing plants, correlating with their increased stress tolerance. Interestingly, 5-6% of leaf-expressed genes were affected by flavodoxin in the absence of drought, representing pathways modulated by chloroplast redox status during normal growth. About 300 of these genes potentially contribute to stress acclimation as their modulation by flavodoxin proceeds in the same direction as their drought response in wild-type plants. Tuber yield losses under chronic water limitation were mitigated in flavodoxin-expressing plants, indicating that the flavoprotein has the potential to improve major agronomic traits in potato.