Environmental and Experimental Botany

Light wavelengths modulate plant growth, metabolism, and physiology. Amaranthus, a C4 underutilized climate resilient crop with promising nutritional properties remained unexplored in terms of metabolite enrichment under monochromatic light wavelengths of visible spectrum. In current study, two cultivars of Amaranthus tricolor (green and red) were exposed to seven light regimes of photosynthetically active radiation (PAR; 400-700 nm): deep blue, blue, green, amber, red, deep red, far red, and their metabolic responses were captured using Gas Chromatography-Mass Spectrometry. The metabolic analysis revealed wavelength-specific reprogramming in the levels of organic acids, sugars, amino acids, fatty acids as well as phenolics. In both the green and red Amaranthus, branched-chain amino acids and phenylalanine, which are nutritionally essential, were significantly elevated under far-red light. While the phenolics such as caffeic acid and ferulic acid were elevated under green and deep blue light respectively in green Amaranthus, amber light wavelengths enhanced these phenolics in red Amaranthus. The study highlighted cultivar-specific metabolic rewiring triggered by specific wavelengths. Altogether, these findings provides insights into metabolic adaptation and demonstrate the ability of light wavelength to specifically enrich the targeted metabolite of nutritional relevance in Amaranthus. It offers strategies to improve the nutritional value of crops in controlled agriculture systems. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=167 HEIGHT=200 SRC="FIGDIR/small/714947v1_ufig1.gif" ALT="Figure 1"> View larger version (40K): org.highwire.dtl.DTLVardef@1a4477dorg.highwire.dtl.DTLVardef@518550org.highwire.dtl.DTLVardef@7682dorg.highwire.dtl.DTLVardef@4876e2_HPS_FORMAT_FIGEXP M_FIG C_FIG

Photosynthesis accounts for most of the final grain yield in rice, making improvements in radiation use efficiency (RUE) a key strategy for enhancing productivity. Agronomically, RUE is defined as the biomass produced per unit of total solar radiation or photosynthetically active radiation intercepted by the canopy. However, the interaction between carbon and nitrogen metabolism plays a critical role in determining plant growth and grain yield. Assimilated nitrogen is required for the synthesis of photosynthetic pigments and enzymes, while the reduction of nitrate (NOLL) and nitrite (NOLL), as well as the assimilation of ammonium (NHLL), depend on the reducing power and carbon skeletons generated by photosynthesis. In this study, two high-yielding rice (Oryza sativa) cultivars--an indica-type (El Paso 144) and a japonica-type (INIA Parao) were subjected to two nitrogen treatments (3 mM and 9 mM NOLL/NHLL) and two light intensities (850 and 1500 mol mL{superscript 2} sL{superscript 1}). A strong interaction between light intensity and nitrogen metabolism was observed, with contrasting responses between subspecies. These differences reflect a coordinated regulation of carbon assimilation and primary nitrogen metabolism. The results provide new insights into the metabolic strategies underlying nitrogen compound accumulation under variable irradiance. Such knowledge is essential for improving nitrogen fertilizer use efficiency and yield performance in elite rice genotypes cultivated under commercial field conditions.

During drought, numerous compounds accumulate in plant tissues, but their physiological roles remain unclear - they may function as osmolytes, osmoprotectants, or merely arise as by-products of stress-induced metabolic shifts. We developed an experimental approach to link accumulation patterns with specific functions, using Scots pine (Pinus sylvestris L.) saplings subjected to water deprivation and subsequent rewatering as a model system. We monitored changes in relative water content (RWC) and osmotic adjustment dynamics, employed untargeted primary metabolite profiling for preliminary screening of compounds correlated with water status, and performed quantitative GC-MS and LC-MS analyses of selected metabolites. Major inorganic cations (K, Ca{superscript 2}, Mg{superscript 2}) were also quantified to assess their potential roles. Our results revealed that tryptophan, valine, and lysine - though generally present in low abundance - exhibited selective accumulation under severely reduced RWC ([≤] 70%), suggesting their involvement as osmoprotectants. Major organic acids, particularly shikimic acid, showed trends consistent with osmotic adjustment. Notably, neither sucrose nor inorganic cations appeared to function as primary osmolytes in this context. The proposed approach offers a viable strategy for identifying compounds involved in plant adaptation to water deficit, with potential applications in breeding programs aimed at improving drought tolerance. HighlightsAn approach to identify osmolytes and osmoprotectants was implemented Accumulation of Trp, Val and Lys was consistent with their role in osmoprotection Osmotic adjustment relied predominantly on organic acids than on inorganic ions Monosaccharides but not sucrose correlates with changes in needle water status

Napier grass (Cenchrus purpureus syn. Pennisetum purpureum), a perennial C4 forage and bioenergy crop, exhibits strong drought resilience, yet the integrative mechanisms underlying this tolerance remain incompletely understood. This study examined physiological, hydraulic, and metabolic responses of four Napier grass cultivars under PEG-induced osmotic stress and progressive soil water deficit. Drought significantly increased the root-to-shoot ratio, indicating preferential biomass allocation to roots, which supported maintenance of shoot growth and tissue water status. All cultivars showed an approximate twofold increase in water-use efficiency (WUE) under water deficit, with cv2 and cv7 displaying superior performance. Upregulation of aquaporin genes (PIP2;2 and PIP2;3) suggested active hydraulic regulation that sustained carbon assimilation under reduced transpiration. Metabolic profiling revealed pronounced root-centered osmotic adjustment, including accumulation of galactinol, myo-inositol, raffinose family oligosaccharides, proline, and several amino acids. Enhanced expression of the galactinol synthase gene confirmed activation of raffinose biosynthesis pathways. Genotypic variation highlighted cv2 as particularly drought resilient. Rapid post-stress regrowth further underscored the importance of perennial root persistence. In conclusion, drought tolerance in Napier grass arises from coordinated hydraulic resilience, osmotic adjustment, and C4 photosynthetic efficiency, supporting its suitability for forage and bioenergy production in water-limited environments. SignificantThis study shows drought tolerance in Napier grass relies on root-driven hydraulic and metabolic regulation with efficient water-use efficiency, rather than avoidance, and that PEG responses predict field performance.

Salinity and sodicity stresses adversely affect rice growth and yield. To overcome yield losses, suitable tolerant rice cultivars can be developed through a marker-assisted breeding (MAB) program. In the present study, genomic regions associated with sodicity stress tolerance at the reproductive stage were identified using a high-density 50kSNP array in a recombinant inbred line (RIL) population derived from the contrasting rice genotypes CSR11 and MI48. A total of 50 QTLs were detected for various yield-related traits; further, 19 QTLs with [≥]15% of phenotypic variance were selected for integrated (omics) analysis. RNA sequencing of leaves and panicles at the reproductive stage under sodic stress conditions was employed to find differentially expressed genes. A total of 1368 and 1410 SNPs; 104 and 144 indels were found for MI48 and CSR11, respectively, within the QTL regions from resequencing. At chromosomes 1 and 6, colocalized QTLs (qPH1-1/qGP1-1 and qGP6-2/qSSI6-2) were discovered. Differentially expressed genes (DEGs) were mapped over the QTL regions selected, and SNP variations and indels were screened for colocalized QTLs. Potential candidate genes, namely Os-pGlcT1 (Os01g0133400), OsHKT2;1 (Os06g0701600) and OsHKT2;4 (Os06g0701700), OsANTH12 (Os06g0699800), and OsPTR2 (Os06g0706400), were identified as being responsible for glucose transport, ion homeostasis, pollen germination, and nitrogen use efficiency, respectively, under salt stress. Finally, our study provides important insights into the genes and potential mechanisms affecting grain yield under sodic stress in rice, which will contribute to the development of molecular markers for rice breeding programs.

The RETINOBLASTOMA-RELATED (RBR) protein in plants functions as a cell-cycle inhibitor, regulating cell numbers in developing organs and establishing cellular quiescence during growth. Although the role of RBR counterparts in animals also involves regulating cell size, this potential function remains unexplored in plants. We investigated transgenic Arabidopsis plants with altered RBR levels and observed corresponding changes in cell size from embryogenesis through organ development. In addition, stomatal meristemoid cells with reduced RBR levels divided beyond the size threshold, whereas elevated RBR levels increased their size. RBR stimulated terminal differentiation in the stomatal lineage by inducing MUTE and CYCLIN D5;1 expression, whereas reduced RBR levels maintained asymmetric divisions through high SPEECHLESS and CYCLIN D3;1 expression. Interestingly, the cell proliferation-dependent phosphorylation of RBR at the conserved 911Ser site positively correlated with RBR protein levels in the transgenic lines and aligned with the effect of RBR on cell size. This study discusses the potential link between RBRs control of cell proliferation and cell size, providing new insights into the coordinated regulation of plant development.

Photosynthetic electron transport is mediated by several protein supercomplexes that are spatially arranged in the thylakoid membranes of chloroplasts. The chloroplast NADH dehydrogenase-like (NDH) complex is part of the photosynthetic alternative electron transport (AET) chain, which reduces the plastoquinone (PQ) pool using reduced ferredoxin as a substrate. This NDH complex is associated with photosystem I (PSI) and mediates a portion of AET in stroma lamellae, whereas photosystem II (PSII) is concentrated in grana stacks. This study presents the findings regarding post-illumination chlorophyll fluorescence increase (PIFI), a protein crucial for regulating AET via the NDH pathway. A marked increase in NDH activity and a reduction in the PQ pool in the dark were observed in PIFI-deficient mutant strains (g-pifi) generated by genome editing. Blue native PAGE analysis indicated that PIFI was associated with the NDH-PSI supercomplex in the wild type, and the NDH complex was dissociated from PSI in the g-pifi mutants. Additionally, the g-pifi mutants exhibited a decrease in the maximum quantum yield of PSII (Fv/Fm). Notably, Fv/Fm was restored in a double mutant harboring both g-pifi and NDH-deficient pnsl1 mutations, demonstrating that deregulated NDH activity in g-pifi causes downregulation of PSII efficiency. However, the lower Fv/Fm was not observed in a mutant lacking thioredoxin m4 (trxm4), which showed deregulated NDH activity but maintained the NDH-PSI supercomplex. These data suggest that PIFI stabilizes the NDH-PSI supercomplex and maintains the spatial localization of PQ reduction via AET in thylakoid membranes, which is essential for the proper functioning of PSII.

Previously, the chitin receptor-interacting protein kinase LIK1 (LysM receptor kinase 1/CERK1-interacting kinase) was shown to play an important role in regulating chitin signaling and plant defense. A limited proteolysis proteomics study revealed several LIK1-derived peptides that showed differential abundance between ATP-treated and mock-treated Arabidopsis samples, suggesting a possible involvement of LIK1 in extracellular ATP (eATP) signaling. To explore this possibility, LIK1 mutants were obtained and examined for their response to ATP. The results showed that mutations in LIK1 significantly reduced the expression of eATP-responsive genes. In addition, LIK1 was found to interact with the eATP receptor P2K1 and to be phosphorylated by it. The LIK1 protein was localized to the plasma membrane and its gene expression appeared to be ubiquitous. Collectively, these findings indicate that LIK1 not only contributes to chitin signaling but also participates in eATP signaling, highlighting its potential role as a shared component in multiple signaling pathways to regulate plant responses to diverse internal and external cues.

AO_SCPLOWBSTRACTC_SCPLOWNitrogen fertilizers are essential for crop productivity but cause environmental harm, necessitating the development of cultivars that thrive under limited nitrogen. This study investigates the transcriptomic response to nitrate in Arabidopsis thaliana (a model dicot), Brachypodium distachyon (a model Pooideae), and Hordeum vulgare (barley, a domesticated Pooideae) to identify conserved and species-specific molecular mechanisms. Using RNA-seq after 1.5 and 3 hours of nitrate treatment, we found that core nitrate-responsive biological processes - such as nitrate transport, assimilation, carbon metabolism, and hormone signaling - are largely conserved across species. However, comparative analysis at gene level based on orthology revealed specificities between the species. For instance, rRNA processing was uniquely stimulated in Arabidopsis, while cysteine biosynthesis from serine and gibberellin biosynthesis were specifically regulated in Brachypodium and barley. Orthologs of key nitrate-responsive genes (e.g., NRT, NLP, TCP20) exhibited variable regulation, reflecting potential adaptations linked to domestication or nutrient acquisition strategies. These findings highlight the importance of integrating model and crop species to uncover targets for improving nitrogen use efficiency in cereals. The study provides a pipeline integrating gene ontology and orthology analyses to compare transcriptomic responses between species.

Improving photosynthesis is a promising approach to enhance sugar beet productivity. However, genetic variation in leaf photosynthesis and its relationship with disease resistance remain underexplored. We evaluated 98 sugar beet genotypes representing different breeding categories, including commercial F1 hybrids, seed-parent lines, and pollinator lines, in Hokkaido, northern Japan. Leaf gas exchange was measured during early growth under field conditions around the infection period of Cercospora leaf spot (CLS). To account for fluctuating irradiance during large-scale phenotyping, we applied a multilevel mixed-effects light-response model to estimate genotype-specific photosynthetic characteristics. Substantial genotypic variations in photosynthetic characteristics were detected. F1 hybrids exhibited higher photosynthetic capacity than breeding lines, whereas differences among breeding categories were unclear due to large within-category variation. Some breeding lines exhibited photosynthetic rates higher than those of hybrids, indicating exploitable genetic resources within the present genetic panel. We did not detect statistically significant trade-off between leaf photosynthesis and CLS resistance among 98 genotypes; in a subset of 19 genotypes analysed in detail, the relationship was even synergistic. Our results highlight the genetic diversity of leaf photosynthesis and its category-dependent structure, and suggest that selection for enhanced photosynthesis can proceed without substantial trade-off with CLS resistance. HighlightLeaf photosynthesis of 98 sugar beet genotypes showed significant genetic variation and dependence on breeding category. Active photosynthesis incurred minimal trade-off with Cercospora leaf spot resistance.

Belowground, plants are exposed to a wide range of biotic stresses that vary in severity and nature, including tissue damage, disruption of vascular connectivity, and depletion of assimilates. How plants adapt their root systems to cope with different types of belowground biotic stresses is not well known. In this paper we compare above- and belowground plant adaptations to three nematode species with distinct tissue migration and feeding behaviours to study mechanisms underlying tolerance to different types of biotic stresses. We monitored both green canopy growth and changes in root system architecture of Arabidopsis inoculated with Pratylenchus penetrans, Heterodera schachtii, and Meloidogyne incognita. This revealed three distinct phases in aboveground plant responses: (i) initial growth inhibition associated with host invasion and tissue damage, (ii) persistent growth reduction associated with nematode sedentarism, and (iii) late growth stimulus in more advanced stages of infection. Specific adaptations in the root systems further revealed fundamentally different stress coping strategies. Tissue damage and intermittent feeding by P. penetrans in the root cortex did not induce significant changes in root system architecture. Tissue damage to the root cortex and prolonged feeding on host vascular cells by H. schachtii induced secondary root formation compensating for primary root growth inhibition. Prolonged feeding on host vascular cell by M. incognita alone did not induce secondary root formation, but was accompanied by typical local tissue swelling instead. Our data suggest that local secondary root formation and tissue swelling are two distinct compensatory mechanisms underlying tolerance to sedentarism by root-feeding nematodes. HighlightHow plants utilize root system plasticity to cope with different types of biotic stresses by root feeding nematodes remains largely unknown. Here, we report on specific adaptive growth responses in Arabidopsis roots to three nematode species, Pratylenchus penetrans, Heterodera schachtii, and Meloidogyne incognita, with fundamentally different strategies for host invasion, subsequent migration through host tissue, and feeding on host cells.

Spinach (Spinacia oleraceae) is a principal vegetable crop commercially grown in Controlled Environment Agriculture (CEA). Recent research suggests that root morphological and architectural differences among crop species influence yield, resource use efficiency, and environmental stress tolerance. These root traits may be exploited to increase yield, promote efficient nutrient use, and mitigate environmental stressors. This study measured differences between various spinach cultivars in CEA systems to reveal morphological and anatomical variation. We grew three spinach cultivars with different reported growing rates ( Income, Darkside, and El-Majestic) under NFT hydroponic and substrate-based systems in a controlled greenhouse environment over 45 days with destructive harvests at days 15, 30, and 45. Supplemental light (250 {micro}mol/m2/s) with 12-hour photoperiod and periodic fertigation was used. Harvests included the collection of leaf and root biomass, and scanning of root systems in WinRhizo software, measuring ten variables. On day 45, root cross-sections from orders 1-5 were embedded in JB-4 resin, sectioned, stained, and analyzed for diameter, vasculature, and rhizodermis characteristics. Results indicate that in spinach, differences in root system morphology are linked to cultivation systems over cultivar identity. Vascular and root anatomical alterations are minor compared to morphological differences in response to the cultivation system. Hydroponic-style growth systems are associated with the proliferation of fine-root ideotypes compared with substrate-based conditions. Such findings affirm previous studies, which suggest plastic root morphology in response to growth systems, and may be used to help create more resilient, resource-efficient cultivars. HighlightsO_LIIn spinach, root system morphology differences are linked to cultivation systems. C_LIO_LIRoot vascular and anatomical alterations are minor in response to cultivation system. C_LIO_LIHydroponic growth systems are linked to fine-root ideotype proliferation in spinach. C_LIO_LIFine-root ideotype proliferation may be a breeding target for CEA spinach. C_LI

Soluble N-ethylmaleimide-sensitive factor attachment protein receptors (SNAREs) are essential regulators of plant growth, development, and stress adaptation. In this study, we performed a comprehensive genome-wide identification of SNARE genes in cucumber (Cucumis sativus L.), uncovering 51 putative members designated as CsSNAREs. Phylogenetic analysis confirmed that these genes cluster into five major clades: Qa-CsSNARE (14), Qb-CsSNARE (9), Qc-CsSNARE (10), Qb+c-CsSNARE (3), and R-CsSNARE (15). Bioinformatic analysis of their promoter regions, coupled with expression profiling under diverse abiotic stress conditions, highlighted a heightened responsiveness within the Qa-CsSNARE subfamily. To validate this, we selected representative Qa-CsSNARE genes for quantitative real-time PCR analysis under drought and salt stress. Among these, CsSYP121 was notably induced by salt treatment. We subsequently generated transgenic cucumber lines overexpressing CsSYP121 and challenged them with salinity. Phenotypic assessment, combined with measurements of reactive oxygen species (ROS) accumulation and K+/Na+ ratios, demonstrated that CsSYP121 overexpression (OE) confers enhanced salt tolerance and boosts antioxidant capacity. We propose a model wherein CsSYP121 mitigates ROS-induced cellular damage under salt stress, potentially through promoting K+/Na+ homeostasis, thereby improving plant performance under saline conditions. Our findings identify CsSYP121 as a promising candidate gene for breeding salt-tolerant crops.

Enset (Ensete ventricosum), a multipurpose crop domesticated exclusively in Ethiopia, serves as a staple food for millions of smallholder farmers. It is primarily cultivated as a monocrop in homegardens, leaving it vulnerable to climate change risks. One potential nature-based solution involves agroforestry systems; however, ensets response to canopy cover remains unclear. This study examined how scattered trees in enset farms affected microclimate and enset morpho-physiology in South Ethiopia. Trees significantly modified the microclimate conditions in enset homegardens. The average daily reductions in air, soil surface, and soil temperatures ranged from -0.5 to -1.9 {degrees}C, -0.4 to -2.1 {degrees}C, and +0.4 to -1.0 {degrees}C, respectively. The minimum soil moisture offset ranged from +0.8% to +5.7%. Although the tree identity effect on enset growth was negligible, planting position relative to the overstory trees significantly influenced enset responses. Most morphophysiological traits were higher under tree canopies, with progressively lower values at the edge and outside the tree canopy. In contrast, leaf dry matter content exhibited an inverse trend, aligning with the leaf economics spectrum. These results demonstrate ensets adaptability to canopy shade, suggesting potential for agroforestry expansion. Cultivar-specific shade tolerance and ideal shade levels to maintain enset productivity should be investigated further.

Heat-tolerant rice cultivars are essential for mitigating global warming impacts. Basal anther dehiscence length (BDL) is a promising visible morphological marker for heat tolerance through stable pollination. We investigated the effects of sowing date on anther morphology, pollination, and fertility under controlled high-temperature conditions (35, 37, or 39 {degrees}C at flowering). Three japonica cultivars-- Akitakomachi (early heading), Koshihikari (medium), and Hatsushimo (late)--were sown monthly over 3 months and grown in pots. At heading, the plants were exposed to the temperature treatments for 3 days, and the proportion of florets with [≥]10 germinated pollen grains on the stigma (GP10) and seed set were assessed. Among anther traits, BDL showed the greatest variation, with all cultivars from the second sowing exhibiting the shortest BDL. Analysis of variance revealed significant effects of genotype, sowing date, and their interaction on anther traits and fertility. Regression analysis indicated that fertility was associated with GP10, with BDL contributing significantly to GP10 in the late-heading Hatsushimo, together with maximum temperature at flowering. Thus, both genotype and environment shape anther morphology, pollination, and fertility, indicating that BDL plasticity and genotype-specific environmental responses must be carefully considered when using BDL as a breeding marker for heat tolerance. HighlightVariation in sowing date significantly affects anther morphology and heat tolerance in rice. Genotype-specific responses to the growing environment require careful consideration for reliable breeding assessments.

Smart irrigation management is essential for improving crop resilience under increasing drought frequency driven by climate change. Although satellite-based remote sensing provides valuable tools for monitoring crop water status at large spatial scales, its accuracy is often limited in mountainous and heterogeneous agricultural landscapes. In this study, we investigated drought-induced metabolic responses in potato (Solanum tuberosum L.) to identify biochemical biomarkers that could complement satellite-based irrigation advisories in the mid-Himalayan region of India. A field experiment was conducted using a gradient of soil moisture regimes corresponding to moderate (50% field capacity), critical (25% field capacity), and extreme drought stress (5-8% field capacity). Satellite-derived evapotranspiration-based irrigation advisories were validated against in situ soil moisture measurements, revealing discrepancies attributed to the inability of satellite estimates to capture actual water loss under drought stress conditions, highlighting the need for additional ground-truth biomarkers across heterogeneous field conditions. To capture plant-level physiological responses, untargeted metabolite profiling of potato leaves was performed using gas chromatography-mass spectrometry (GC-MS). Approximately fifty metabolites belonging to amino acids, organic acids, sugars, and sugar alcohols were detected. Multivariate statistical analyses revealed distinct metabolic signatures associated with progressive drought stress. Notably, accumulation of proline, serine, isoleucine, sucrose, fructose, glucose, and polyols such as mannitol and myo-inositol reflected key metabolic reprogramming associated with osmoprotection, redox homeostasis, and energy metabolism under drought conditions. Collectively, this ensemble of stress-responsive metabolites represents a robust panel of drought stress biomarkers. As a proof of concept, proline was validated as a qualitative biomarker of plant water status through a rapid and cost-effective colorimetric biochemical assay, demonstrating its practical applicability for field-level irrigation management. These findings demonstrate that metabolomics-derived biomarkers can provide sensitive plant-level indicators of drought stress that complement satellite-based monitoring systems. The integration of biochemical diagnostics with remote sensing platforms offers a promising approach for improving drought detection and developing low-cost, field-deployable tools for smart irrigation advisories in heterogeneous agricultural landscapes. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=93 SRC="FIGDIR/small/712810v1_ufig1.gif" ALT="Figure 1"> View larger version (33K): org.highwire.dtl.DTLVardef@59919corg.highwire.dtl.DTLVardef@66ce49org.highwire.dtl.DTLVardef@17143dcorg.highwire.dtl.DTLVardef@11e2769_HPS_FORMAT_FIGEXP M_FIG C_FIG

Malate and fumarate constitute a significant transient carbon stock that is dynamically synthesized during the photoperiod. These organic acids are diurnally stored and remobilised from the vacuole, and they have a key role in the cellular metabolic regulation. This function is well known in C4 and CAM plants. However, in C3 species that are the majority of terrestrial plants, the importance of the vacuolar accumulation/release and its influence on plant growth is still an open question. In Here we addressed this issue generating multiple knockout mutants in Arabidopsis thaliana lacking vacuolar anion channels of the Aluminium-Activated Malate Transporter (ALMT) family, to impair malate and fumarate transport to the vacuole. We show that in these mutants reducing vacuolar transport of malate and fumarate in mesophyll cells leads to a dramatic growth impairment. Metabolic and fluxomic analysis revealed that vacuolar malate and fumarate transport influences plant carbon and nitrogen metabolism as well as cellular pH and ionic homeostasis. In conclusion, our results show that the transport organic acids like malate and fumarate across the vacuolar membrane is essential for plant growth in a C3 plant too. These results establish the importance of the vacuolar pools of malate and fumarate in plant metabolism.

The chloroplast NADH dehydrogenase-like (NDH) complex mediates cyclic electron transport (CET) around photosystem I (PSI) and contributes to photosynthetic regulation and photoprotection under various environmental stresses. Although NDH function has been extensively characterized under controlled conditions, NDH-deficient mutants often show only subtle phenotypes in such environments, leaving its physiological importance under naturally fluctuating field conditions poorly understood. Here, we evaluated growth, yield, and photosynthetic performance of NDH-deficient rice cultivated under field conditions. Mutant plants exhibited reduced biomass accumulation and grain yield compared with wild type. Detailed physiological analyses revealed that NDH deficiency markedly decreased PSI electron transport and CO2 assimilation, particularly under low temperature and sub-saturating irradiance. At moderate and high temperatures, reductions in carbon fixation were largely confined to low-light conditions, whereas at low temperatures, impairment extended across nearly the entire light response range. Under repetitive fluctuating light regimes, NDH-deficient plants showed progressive declines in photosynthesis accompanied by a selective decrease in PSI photochemical capacity without changes in PSII maximum efficiency, indicating PSI-specific photoinhibition. These findings demonstrate that NDH-dependent CET plays a crucial role in sustaining photosynthetic efficiency and crop productivity in dynamic field environments by stabilizing PSI redox balance and maintaining long-term carbon gain. Summary StatementNDH-dependent cyclic electron transport supports photosynthesis and yield in field-grown rice by maintaining PSI function under fluctuating light, low temperature, and sub-saturating irradiance.

Small signaling peptides (SSPs) are critical regulators of plant growth, development, and responses to biotic and abiotic stress, yet their role in the C4 grass Sorghum bicolor is largely uncharacterized. To help fill this knowledge gap, 219 S. bicolor genes that encode SSPs were identified based on SSP sequences previously identified in Arabidopsis thaliana, Oryza sativa, Zea mays, Triticum aestivum, and Brachypodium distachyon. The 219 sorghum genes were assigned to 19 gene families, analyzed for the presence of motifs, and aligned with genes that encode SSPs in other plants using phylogenetic analysis. Expression of the 219 SSP encoding genes in sorghum organs, during stem development, and in stem tissues and cell types revealed distinct spatial, temporal and developmental patterns of expression. Genes associated with the SbCEP and SbRGF families were preferentially expressed in roots, whereas SbEPF genes were expressed in stems and panicles. The expression of genes during bioenergy sorghum stem growth and development was investigated because stems account for [~]80% of harvested biomass and serve as conduits for water and nutrient transport between leaves and roots. During stem development, 28 SSP encoding sorghum genes in several families (CLE, EPF, CEP, GASS, PSY, ES, PSK, CAPE, POE) were expressed at higher levels in zones of cell proliferation. For example, the TDIF homologs SbCLE41 and SbCLE42 were expressed at high levels in nascent stem nodes where they may regulate cambial activity and vascular bundle cell differentiation. A different set of 15 genes in the CIF, POE, CAPE, PSY, CEP, RALF, and CLE families were expressed at higher levels in zones of stem tissue differentiation highlighted by elevated expression of 5 SbRALFs in the stem nodal plexus. Cell type specific expression of many SSP encoding sorghum genes was also observed in fully elongated internodes indicating gene expression is regulated with high spatial resolution. Overall, the results provide a foundation of information for analysis of SSP functions in sorghum that can be integrated with knowledge of sorghum gene regulatory networks to modulate traits important for production of sorghum crops.

Understanding the regulation of enzyme activity involved in photosynthesis is essential for engineering enhanced carbon fixation in crops. In C4 plants, the enzyme phosphoenolpyruvate carboxylase (PEPC, EC 4.1.1.31) is one of the most abundant leaf enzymes and plays an essential role in photosynthetic carbon dioxide (CO2) fixation. The enzyme also plays a key role in central metabolism (e.g., providing intermediates to the citric acid cycle) and therefore must be highly regulated to coordinate its activity. The regulation of PEPC activity can occur allosterically by glucose 6-phosphate activation and malate inhibition, which is in part influenced by reversible phosphorylation. A specific light-dependent phosphorylation of PEPC at an N-terminal serine residue by the PEPC-protein kinase (PEPC-PK) can regulate its sensitivity to this allosteric regulation. However, the impact of this PEPC phosphorylation has not been tested in a C4 crop. Therefore, we created PEPC-PK mutant lines in Zea mays to assess the impact of PEPC phosphorylation on its allosteric regulation, photosynthesis, and growth. While the maximum PEPC activity was unchanged, PEPC in the PEPC-PK mutant plants was not phosphorylated under light and was more sensitive to malate inhibition. However, gas exchange, electron transport, and field biomass analyses showed no differences in the PEPC-PK mutant plants. These results demonstrate that in Z. mays PEPC phosphorylation affects enzyme sensitivity to malate in vitro but does not substantially alert photosynthetic performance or growth under field conditions suggesting additional regulation of PEPC activity in planta.