Plant Physiology and Biochemistry

Blue light using Flavin (BLUF) proteins are microbial photoreceptors that are involved in various physiological responses. Their occurrence and biochemical properties in fungi remain poorly understood. Here, we investigated a putative BLUF photoreceptor from the corn-smut fungus Mycosarcoma maydis (MmBLUF). Domain analysis, multiple sequence alignment of BLUF core regions, and structural modelling indicated conserved canonical BLUF fold and flavin-pocket residues. However, when heterologously expressed, UV-visible and fluorescence spectroscopy revealed different spectral behaviour than canonical BLUF protein. Further, we tested the role of extended N-terminus in modulation of chromophore binding by expressing N-terminus truncated protein variants. Our results suggest that the unusual spectral behaviour is not linked to the truncation construct (extended N-terminus), which also showed similar spectral features, indicating that the extended N-terminus is unlikely to account for an unusual photodynamics characteristics. Our findings support MmBLUF as a structurally conserved putative fungal BLUF-like photoreceptor with different photochemical properties. Further studies are required to establish its chromophore identity, photocycle and function of this unusual BLUF-like domain from fungal system.

Tomato production, yield and fruit quality face major challenges due to several factors, including the complex polygenic inheritance of agronomically relevant traits, biotic and abiotic stresses, and increasingly stringent regulations limiting the use of phytosanitary products. In this context, bioinoculants have emerged as a sustainable strategy capable of enhancing yield without compromising fruit quality, conferring protection against different stresses and exerting a minimal or no impact on environment and human health. In this study, we evaluated the effects and the underlying mechanisms by which Streptomyces sp. N2A, an actinobacteria isolated from soybean rhizosphere, promotes seed germination, vegetative growth and yield in tomato, without modifying fruit quality. The obtained results demonstrated that the bacterial treatment significantly improved seedlin[g]s emergence and growth and development in vegetative stage. At harvest, yield was also significantly enhanced, mainly driven by increased individual fruit weight, which was positively correlated with a thicker pericarp in fruits from N2A-treated plants. Transcriptional analysis during fruit development revealed a coordinated induction of auxin and cytokinin signaling pathways before and after anthesis, providing a hormonal framework that underlies the promotion of pericarp growth. This study provides evidence of the beneficial effect of inoculation with Streptomyces sp. N2A on tomato yield and constitutes the first report describing the modification of fruit morphology and expression of genes involved in phytohormonal modulation during early growth and development, induced by a plant growth-promoting Streptomyces.

The ability of seeds to survive until dormancy recedes and the germination requirements are met is an adaptive strategy. Proteomics improves our understanding of the mechanisms that control the adaptation to environmental heterogeneity. In this study, we investigated two European beech populations from different habitats that differed in dormancy and germination traits. We found that the populations exhibited different germination strategies, which were reflected in coordinated but quantitatively different proteomic reprogramming. The Miekinia population exhibited stronger accumulation of proteins involved in nucleotide sugar biosynthesis, S-adenosylmethionine metabolism, and flavonoid biosynthesis. Enhanced nucleotide sugar biosynthesis indicates more intensive cell wall remodelling and carbohydrate metabolism, which support embryo growth and faster germination. Increased S-adenosylmethionine metabolism suggests the epigenetic and hormonal regulation of germination differences between populations. Higher flavonoid biosynthesis indicates an enhanced antioxidant capacity associated with environmental protection. In contrast, the Wisa population showed stronger accumulation of proteins involved in RNA processing, suggesting tighter post-transcriptional regulation and proteome reorganization during germination. Consistent with its deeper dormancy and later germination, the Wisa population appears to rely more on RNA-level regulation, whereas the Miekinia population prioritizes metabolic activation. These contrasting proteomic profiles likely reflect population-specific physiological strategies associated with dormancy depth and adaptation to different climatic conditions. HighlightProteomic reprogramming reveals population-specific germination strategies in European beech, linking dormancy depth with contrasting metabolic activation and RNA-level regulation during the transition from dormancy to germination.

Utricularia gibba L. is an aquatic carnivorous plant with a diverse set of capabilities. Reddening of traps frequently occurs in old in vitro cultures. While anthocyanins are often responsible for red coloration in plants, not every plant turns red. Stress factors like high light or excess sucrose have previously been shown to induce the formation of anthocyanins. Here, we hypothesized the red trap formation to be dependent on nutrient deprivation and tested nitrogen deprivation. The results suggest, that only in combination with light, nitrogen deficiency leads to the activation of the complete anthocyanin biosynthesis pathway and visible red coloration. However, in darkness, anthocyanin biosynthesis appears generally less active compared to light conditions and expression of most anthocyanin biosynthesis genes is not significantly upregulated under nitrogen deficiency.

Soybean, a widely cultivated leguminous crop valued for its protein, amino acids, and oil, faces the challenge of maintaining protein levels, which have an inverse correlation with yield. Reducing leaf chlorophyll levels could increase seed protein levels without compromising yield; however, this is yet to be tested. Therefore, to understand the impacts of low chlorophyll mutations on soybean yield and seed composition, we screened and compared 25 low chlorophyll soybean mutants to their 11 dark green parents. PI548210 (Lincoln mutant) demonstrates a higher concentration of protein without affecting yield compared to its dark green parent PI548362 (Lincoln), suggesting it as a good candidate for further large-scale field trials. PI547555 (Y11/y11, Clark mutant) demonstrates a lower concentration of oil without impacting yield, alongside lower gross photosynthesis, but with chlorophyll levels in the pod and seed tissues that are comparable to its dark green parent PI548533 (Clark). These findings are consistent with the oil concentration of the soybean being influenced by pod and seed photosynthesis, which is correlated with pod height and row spacing. Chlorophyll levels in the leaf do not necessarily correlate with those in the pod and seed of low chlorophyll mutants, possibly due to substantially lower expression of chlorophyll synthesis genes in the pod and seed. SIGNIFICANCEO_LIPI548210 (Lincoln mutant), one of twenty-five low chlorophyll soybean mutants, demonstrates a higher concentration of soybean protein without affecting yield compared to its dark green parent (Figure 1 and Table 1). C_LIO_LIPI547555 (Y11/y11, Clark mutant), a low chlorophyll soybean mutant, demonstrates a reduced concentration of soybean oil without impacting yield, alongside lower gross photosynthesis in pod and seed tissues compared to its dark green parent (Figures 3 and Table 2). These findings suggest that the oil concentration of the soybean is influenced by pod and seed photosynthesis, which is in turn influenced by pod height and row spacing (Figure 2). C_LIO_LIChlorophyll levels in the leaf do not necessarily correlate with those in the pod and seed of low chlorophyll mutants, possibly due to substantially lower expression of chlorophyll synthesis genes in the pod and seed (Figure 5-6). C_LI O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=84 SRC="FIGDIR/small/744892v1_fig1.gif" ALT="Figure 1"> View larger version (55K): org.highwire.dtl.DTLVardef@4282dcorg.highwire.dtl.DTLVardef@9d565forg.highwire.dtl.DTLVardef@1918292org.highwire.dtl.DTLVardef@1359b1_HPS_FORMAT_FIGEXP M_FIG O_FLOATNOFigure 1.C_FLOATNO Two low chlorophyll mutants are as healthy as their dark green parents. Lincoln and its low chlorophyll mutant, left; Clark and its low chlorophyll mutant, known as Y11/y11, right. It can be seen by eye that the plants have low chlorophyll (light green/yellow leaves) but a similar growth habit to their dark green parents. See Supplemental Figures 1-4 for contrast, where low chlorophyll mutants are stunted in growth compared to their dark green parents. C_FIG O_TBL View this table: org.highwire.dtl.DTLVardef@657ec9org.highwire.dtl.DTLVardef@166e75borg.highwire.dtl.DTLVardef@df23c7org.highwire.dtl.DTLVardef@1a60124org.highwire.dtl.DTLVardef@194ed96_HPS_FORMAT_FIGEXP M_TBL O_FLOATNOTable 1.C_FLOATNO O_TABLECAPTIONComparison of seed yield, weight, seed composition between low chlorophyll mutants and their dark green parents. ANOVA is used with linear mixed model (random effect = block, fixed effect = variety). Least squares mean is used to compare. For yield and seed composition, N=4 blocks. For leaf chlorophyll (SPAD), N=40. Yield is average yield per plant (g). n.s. = not significant. C_TABLECAPTION C_TBL O_FIG O_LINKSMALLFIG WIDTH=179 HEIGHT=200 SRC="FIGDIR/small/744892v1_fig3.gif" ALT="Figure 3"> View larger version (26K): org.highwire.dtl.DTLVardef@7a368aorg.highwire.dtl.DTLVardef@192b8f0org.highwire.dtl.DTLVardef@1abb738org.highwire.dtl.DTLVardef@89e978_HPS_FORMAT_FIGEXP M_FIG O_FLOATNOFigure 3.C_FLOATNO Light response curve of low chlorophyll mutant (Y11/y11, PI547555) and its parent (Clark, PI548533). Rates of net and gross photosynthesis of low chlorophyll (white) and dark green parents (black) pods under field conditions. Each dot represents a value (n=4) {+/-}SE. We assumed that the seeds greatly inhibited the transmittance of light through the pod and used photosynthetic photon flux density for a single-side. C_FIG O_TBL View this table: org.highwire.dtl.DTLVardef@3f0528org.highwire.dtl.DTLVardef@16ba712org.highwire.dtl.DTLVardef@a5ab2aorg.highwire.dtl.DTLVardef@889254org.highwire.dtl.DTLVardef@3efa4f_HPS_FORMAT_FIGEXP M_TBL O_FLOATNOTable 2.C_FLOATNO O_TABLECAPTIONPod photosynthetic parameters for low chlorophyll mutant (Y11/y11, PI547555) and its parent (Clark, PI548533). Photosynthesis was measured 1 September through 15 September 2021 at the University of Illinois Energy Farm in Urbana, IL, USA. The statistical analysis was done using ANOVA with linear mixed model (alpha=0.05). N=4 {+/-} SEM for Clark and N=3 {+/-} SEM for Y11. C_TABLECAPTION C_TBL O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=130 SRC="FIGDIR/small/744892v1_fig2.gif" ALT="Figure 2"> View larger version (23K): org.highwire.dtl.DTLVardef@a36c26org.highwire.dtl.DTLVardef@1116c8forg.highwire.dtl.DTLVardef@ee5e61org.highwire.dtl.DTLVardef@1766712_HPS_FORMAT_FIGEXP M_FIG O_FLOATNOFigure 2.C_FLOATNO Low chlorophyll mutant (Y11/y11, PI547555) and its parent (Clark, PI548533) differ in concentration of seed oil, which interacts with height of pod and row spacing. The box plots show the median (central line), the lower and upper quartiles (box) and the minimum and maximum values (whiskers). The statistical analysis was done using ANOVA with linear mixed model (n=3 blocks, alpha=0.05). Least squares mean is used to compare. N.s., non- significant in the analysis. A. Concentration of oil in low chlorophyll mutant seeds from the upper canopy decreased by 4% compared to the dark green parent (18.2% vs 19%) while there was no difference between them in the seeds from the lower canopy (20.2% vs 20.6%). B. Schematic layout of 2013 field setting showing two different row spacings. C. Concentration of oil in low chlorophyll mutant decreased by 2% in 38cm spacing (21.4% vs 22%) while there was no difference in 19cm spacing (21.3% vs 21.7%) in 2013 field. C_FIG O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=162 SRC="FIGDIR/small/744892v1_fig5.gif" ALT="Figure 5"> View larger version (22K): org.highwire.dtl.DTLVardef@68e508org.highwire.dtl.DTLVardef@94a6ccorg.highwire.dtl.DTLVardef@152a187org.highwire.dtl.DTLVardef@1eae137_HPS_FORMAT_FIGEXP M_FIG O_FLOATNOFigure 5C_FLOATNO (greenhouse). Correlation between the level of leaf chlorophyll (x-axis: SPAD reading) and the level of immature pod or seed chlorophyll (y-axis, mg/g DW). Line represents the linear regression model. R-squared is a coefficient of determination, the percentage of the response variable variation that is explained by the linear model. Pod is labeled by the fresh weight of seeds it contained. A. Level of chlorophyll of 25-100mg pod (n=18). B. Level of chlorophyll of 100-200mg pod (n=17) . C. Level of chlorophyll of 25-100mg seed (n=17). D. Level of chlorophyll of 100-200mg seed (n=20). C_FIG O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=180 SRC="FIGDIR/small/744892v1_fig6.gif" ALT="Figure 6"> View larger version (28K): org.highwire.dtl.DTLVardef@167fd88org.highwire.dtl.DTLVardef@361472org.highwire.dtl.DTLVardef@786325org.highwire.dtl.DTLVardef@1b53855_HPS_FORMAT_FIGEXP M_FIG O_FLOATNOFigure 6.C_FLOATNO Levels of gene expression in chlorophyll synthesis pathway. A. CHL common pathway genes; Glutamyl-tRNA reductase (GluTR). Glutamate 1- semialdehyde aminotransferase (GSA-AT). ALA dehydratase (ALAD). Uroporphyrinogen III synthase (UROS). Uroporphyrinogen III decarboxylase (UROD). Protoporphyrinogen IX oxidase (PPO). B. Mg branch; Mg-chelatase (Mgch). Magnesium-protoporphyrin IX monomethyl ester cyclase (MPEC). Protochlorophyllide reductase (POR). 3,8-divinyl protochlorophyllide a 8-vinyl-reductase (4VCR). Heme pathway; Ferrochelatase (FECH). Heme oxygenase (HO). Phytochromobilin synthase (HY). Data come from Severin et al (2010). RPKM, reads per kilobase per million mapped reads. DAF, days after flowering. The source seed is experimental line A81-356022 which was generated by introgressing G. soja (PI468916) into G. max (A81-356022). C_FIG

Stomata are the pores on plant surface, and these tiny pores are responsible for the flow of gas between plants and atmosphere. Currently, what effects of the broad and continuous increase in stomatal density achieved via genetic engineering on plant growth and development remain poorly understood. The 9 Arabidopsis transgenic lines with increased stomatal density were acquired through overexpressing FSTOMAGEN (the homologs of STOMAGEN, which are in Flaveria). The intermediate stomatal density (SD) lines exhibited increased trend in biomass. Compared with the lines with low SD, the biomass of Arabidopsis lines with intermediate SD (484 mm-2) significantly increased. There was a positive and significant correlation between biomass and relative water content. Across these transgenic lines, only during the earlier phase of growth, the leaf area exhibited a gradually increased trend as stomatal density increased, and there was both a significant linear relationship between SD and leaf growth rate and a strong linear relationship between SD and leaf area. In contrast, a clear relationship during the later phase wasnt observed. Under lower growth light intensity, there was an increased trend of biomass from other lines to the lines with intermediate SD, and the photosynthetic rate and stomatal conductance of the intermediate line were significantly increased. This study reveals plant-growth alterations that correspond to broad and near-continuous increases in stomatal density achieved via genetic engineering. Our study sheds light on the prerequisites for elevated stomatal density achieved via genetic engineering to promote plant growth.

Accelerating photoprotective regulation to improve carbon assimilation is a promising strategy to increase crop productivity. Although rapid non-photochemical quenching (NPQ) relaxation has been validated as a target through metabolic engineering, it remains unclear whether conventional breeding has improved this trait. Here, we investigated whether more than a century of soybean breeding enhanced NPQ relaxation alongside light-saturated carbon assimilation and seed traits. We evaluated a historical panel of 24 soybean genotypes across vegetative and reproductive developmental stages by integrating NPQ relaxation, gas exchange parameters, xanthophyll-cycle pigment profiles, expression of key photoprotective genes (VDE, PsbS, and ZEP), seed number and seed weight. NPQ relaxation parameters were not consistently associated with genotype release year, seed number, or seed weight at either developmental stage. The only exception was the amplitude of the rapidly relaxing NPQ component (AqE), which was negatively correlated with all three variables during the reproductive stage. In contrast, genotype release year was positively associated with maximum net CO2 assimilation rate (Amax), maximum carboxylation rate of Rubisco (Vcmax), maximum electron transport rate (Jmax), seed number, and seed weight, while Amax and Vcmax were positively correlated with seed number and seed weight. These findings indicate that the greater photosynthetic capacity of modern genotypes was not accompanied by faster photoprotective response. Thus, photoprotective regulation has not kept pace with gains in photosynthetic capacity under field conditions. We conclude that rapid NPQ relaxation remains an important target for synchronizing photoprotection with the high photosynthetic capacity of modern soybean lines.

Chloroplasts comprise photosynthesis and other important processes. Plants protect chloroplasts from stresses including Cd accumulation. Common terrestrial plants, excluders apply a set of mechanisms to restrict Cd penetration to chloroplasts. Removal of accumulated Cd from chloroplasts should also be a beneficial strategy. However, we do not know whether excluder plant species have ability to remove Cd from chloroplasts. We used barley as a common excluder plant species. To barley plants, we applied a model with two stable isotopes 111Cd and 114Cd to distinguish Cd accumulated earlier and later. A portion of Cd absorbed by roots continued translocation to shoot for some days after the external source of Cd was changed from one isotope to another. Chloroplasts acquired new portions of Cd and lost part of Cd accumulated earlier; a total Cd content remained rather unchanged. Cd loss from thylakoids was detected in vivo and in vitro. Cd loss from stroma and envelope was observed in vivo but not in vitro. Therefore, barley has at least two distinct mechanisms for Cd removal from chloroplasts: one from thylakoids and another from stroma. We hypothesized diverse chlorophagy pathways as a potential mechanism for Cd removal from chloroplasts. Cd accumulation by chloroplasts was mainly light-independent. In chloroplasts, Cd accumulated in vivo was tightly bound and mainly located in thylakoids. In vitro, chloroplasts from Cd-treated plants accumulated much less Cd than chloroplasts from untreated plants in a previous study. This implies reorganization of transport across chloroplast envelope membranes. HighlightsO_LICd was removed from thylakoids both in vivo and in vitro C_LIO_LICd was removed from stroma and envelope in vivo but not in vitro C_LIO_LIIn chloroplasts, Cd accumulated in vivo was tightly bound C_LIO_LICd accumulation by chloroplasts was mainly light-independent C_LIO_LIRoot barrier slowed down Cd translocation to shoot but not halted it C_LI

Aquaporins at the plant plasmalemma are divided into two highly conserved subclasses, PLASMA MEMBRANE INTINSIC PROTEINs 1 (PIP1) and PIP2. Arabidopsis thaliana encodes five PIP1 and eight PIP2 isoforms. Individual loss-of-function mutants had been employed for functional analyses. Here, we observe that the pip2;1 pip2;2 pip2;4 pip2;6 pip2;7 quintuple mutant defective of major PIP2 isoforms concomitantly leads to a strongly reduced PIP1 protein level. Lower order mutants pip2;1 pip2;2 and pip2;1 pip2;2 pip2;7 still harbor only 60% and 20% residual PIP1, respectively. This repression is established post-translationally, since neither PIP1s steady-state transcripts nor polysome-associated PIP1 mRNAs are suppressed by pip2;1 pip2;2 pip2;7. Thus, the two major pathways operating in eukaryotes for removal of aberrant proteins, ubiquitin proteasome system (UPS)-dependent ER-associated degradation (ERAD) and autophagy/vacuole-linked degradation, were assessed. Introgression of atg7 blocking autophagy-mediated degradation does not affect the PIP1 protein level of pip2;1 pip2;2 pip2;7. In contrast, introgression of ERAD loss-of-function mutations hrd1A hrd1B and dln1 into pip2;1 pip2;2 pip2;7 partially stabilizes its PIP1 protein level. PIP1 accumulates intracellularly upon pharmacological inhibition of proteasomal degradation by MG132. Nevertheless, the lack of a full PIP1 recovery by these means suggests the flexible operation of parallel ERAD components or unknown pathways. In conclusion, the essential dependence of PIP1 expression on PIP2 isoforms intrinsically interconnects the two PIP subclades at the protein level and will thereby affect their mutual functions. Significance statementPlasma membrane intrinsic proteins constituting the most homogenous plant aquaporin family are nonetheless split into two highly conserved subfamilies, PIP1 and PIP2. The loss of major Arabidopsis PIP2 isoforms does not lead to compensation by PIP1 members, but rather to PIP1s concomitant, post-translational repression. This dependence of PIP1 isoforms inevitably ties the two PIP subfamilies and their function.

Potato (Solanum tuberosum L.), the worlds fourth most cultivated crop, suffers yield losses of up to 40-50% from early blight caused by the necrotrophic fungal pathogen Alternaria solani. In this study we performed gas chromatography-mass spectrometry (GC-MS)-based untargeted metabolomics to characterize temporal alterations in metabolite composition, metabolic pathway regulation, and discriminatory biomarker metabolites in the susceptible Indian potato variety Kufri Jyoti, analyzing infected leaves, non-infected leaves, and lesion-associated necrotic tissues across four days post-inoculation (DPI).Metabolite annotation identified 58 compounds, including sugars, organic acids, amino acids, and secondary metabolites.. Multivariate analyses resolved distinct, largely non-overlapping metabolic clusters for control, infected leaves (1-4 DPI), and lesion tissue (Bs1-Bs3). A biphasic metabolic response was observed: early infection (1-2 DPI) was characterized by general suppression of primary metabolism, while late infection (3-4 DPI) showed pronounced upregulation of glycolysis, the TCA cycle, GS/GOGAT, and the shikimate pathway. Key discriminatory metabolites included asparagine, oxoproline, GABA, phenylalanine, and aromatic amino acids. Lesion tissues exhibited distinct metabolic fingerprints, with early disruption of amino acid recycling followed by a late rebound of defense-associated metabolites. Notably, defence-associated phenolics were detected exclusively within lesion tissue and were absent from whole-leaf profiles, demonstrating that spatially resolved lesion sampling captures defence chemistry that whole-leaf analysis alone would miss. The identified biomarker metabolites, particularly those linked to the shikimate and GS/GOGAT pathways, represent promising candidates for metabolite-assisted breeding and targeted crop protection strategies against early blight in potato. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=115 SRC="FIGDIR/small/745268v1_ufig1.gif" ALT="Figure 1"> View larger version (36K): org.highwire.dtl.DTLVardef@18131edorg.highwire.dtl.DTLVardef@f4fbe6org.highwire.dtl.DTLVardef@1c5db61org.highwire.dtl.DTLVardef@c5ef6d_HPS_FORMAT_FIGEXP M_FIG C_FIG

Background Strigolactones (SLs) regulate diverse aspects of plant development and have been implicated in promoting leaf senescence. However, senescence phenotypes associated with SL deficiency have not been consistently observed across species, suggesting that this function may be species- or context-dependent. Moreover, the contribution of endogenous SL biosynthesis to senescence in leafy vegetable crops remains unclear. Here, we investigated the role of the SL biosynthetic gene MORE AXILLARY GROWTH1 (MAX1) in dark-induced leaf senescence in lettuce (Lactuca sativa). Results We found that endogenous SL biosynthesis plays a major role in dark-induced senescence in lettuce. SL pathway genes were induced during dark storage, while exogenous GR24 accelerated senescence and lettuce MAX1 (LsMAX1) complemented the delayed-senescence phenotype of the Arabidopsis max1 mutant. Consistent with these findings, CRISPR/Cas9-generated Lsmax1 mutants exhibited a pronounced stay-green phenotype during prolonged darkness, accompanied by strongly reduced induction of key senescence-associated genes. Despite this delayed visible senescence, Lsmax1 retained a substantial transcriptional response to dark storage. Strikingly, loss of LsMAX1 did not simply weaken the wild-type senescence program, but redirected part of the response toward a distinct stress-associated transcriptional state that was largely absent from wild type. Loss of LsMAX1 did not affect vegetative rosette architecture, although increased branching emerged after bolting. Conclusions Our findings establish MAX1-dependent SL biosynthesis as an important regulator of leaf senescence in lettuce and reveal a role that extends beyond controlling the rate of senescence. Rather than simply delaying the wild-type program, loss of LsMAX1 alters the transcriptional trajectory of senescence, favoring an alternative stress-associated state during prolonged darkness. The strong stay-green phenotype without detectable changes to vegetative rosette architecture further highlights SL biosynthesis as a potential target for extending postharvest longevity in lettuce and other leafy crops.

Non-photochemical quenching (NPQ) dissipates excess absorbed light energy and protects photosynthetic organisms from photodamage, but its role in regulating the balance between growth, stress tolerance and astaxanthin accumulation in Haematococcus lacustris remains unclear. Here, we investigated how enhanced NPQ affects photosynthetic performance, stress-induced differentiation, and productivity in this astaxanthin-producing microalga. We isolated and characterized an NPQ-enhanced mutant line, A116, using cultivation assays under different stress conditions, analysis of photosynthetic parameters, pigment profiling, and whole-genome resequencing. A116 displayed stronger and faster NPQ induction, driven by increased LHCSR accumulation, resulting in decreased photosynthetic electron transport and lower photochemical efficiency under moderate-to-high light. Enhanced NPQ delayed the transition to astaxanthin-rich cysts under high light, allowing greater biomass accumulation under CO2-limiting conditions. However, under high CO2 availability, where carbon fixation relieved excitation pressure supporting efficient photosynthesis, the enhanced NPQ phenotype reduced growth and astaxanthin productivity compared with the wild type. These results show that NPQ modulates a context-dependent trade-off between photoprotection and productivity in Haematococcus lacustris. Increased energy dissipation can improve high-light tolerance under carbon limitation, but becomes detrimental when absorbed light can be efficiently used for carbon assimilation. Thus, optimal algal productivity requires tuning photoprotective capacity to environmental conditions rather than maximizing NPQ.

Tropospheric ozone (O) is a major atmospheric pollutant that affects plant carbon metabolism, redox homeostasis, and secondary metabolism, including the biosynthesis and emission of biogenic volatile organic compounds (BVOCs). However, the contribution of BVOCs to O3 tolerance, particularly in tropical woody species, remains poorly understood. Here, we investigated whether acute O exposure (cumulative AOT40 of 3497.82 ppb h) induces alterations in photosynthetic performance, redox homeostasis, and BVOC partitioning in Eugenia uniflora. We evaluated gas exchange, photosynthetic pigments, ascorbate and glutathione pools, emitted BVOCs, modeled intercellular BVOC concentrations, and the relative carbon cost associated with BVOC emissions. O exposure significantly increased net CO2 assimilation without affecting stomatal conductance, transpiration, leaf water status, or chlorophyll concentrations, indicating maintenance of photosynthetic performance. Carotenoid concentrations and total glutathione decreased, whereas glutathione redox status was maintained. O induced marked compound-specific changes in BVOC composition and partitioning. Several monoterpenes appeared exclusively under O exposure, {gamma}-elemene emission increased significantly, and the relative distribution of individual BVOCs between the modeled intercellular and emitted pools was altered. These findings show that the response of E. uniflora to acute O exposure was characterized by interplay among carbon assimilation, glutathione redox regulation, and BVOC partitioning rather than by increased total volatile emission. Enhanced carbon assimilation occurred without additional carbon loss through BVOC release, while changes in the modeled intercellular pool indicate that part of the volatile response remained within the leaf. Our findings highlight BVOC partitioning as an important dimension of the plant response to oxidative stress and demonstrate that emission measurements alone may not fully capture the fate and potential physiological role of volatile carbon under O exposure. O_FIG O_LINKSMALLFIG WIDTH=178 HEIGHT=200 SRC="FIGDIR/small/743946v1_ufig1.gif" ALT="Figure 1"> View larger version (52K): org.highwire.dtl.DTLVardef@653af1org.highwire.dtl.DTLVardef@ca5forg.highwire.dtl.DTLVardef@1e641bforg.highwire.dtl.DTLVardef@1e68fae_HPS_FORMAT_FIGEXP M_FIG C_FIG BVOC Partitioning Contributes to Oxidative Stress Defence Under Acute O Exposure

Context. Barley production requires advanced knowledge of its response to changing environmental conditions in order to keep it competitive and sustainable. Aim. Advance in the understanding of barley phenology and foliar development under South American field conditions. Methods. 8 spring barley genotypes with differential phenology were studied in four field experiments under different temperature (through years and sowing dates) and photoperiod (through sowing dates) conditions. Time to anthesis, emergence to onset of stem elongation, stem elongation to anthesis, photoperiod response (PR) in these three traits, number of final leaves at anthesis (FLN) and phyllochron were measured. Key Results. Time to anthesis and its subphases were shorter in late plantings but under similar photoperiod, temperature increased them. Cultivars have differential responses but with magnitude interactions and not crossover ones. Cultivar effects defined PR with no interaction with year (temperature). Temperature and photoperiod affected FLN, phyllochron and their relationship with time to anthesis. Under the shorter photoperiod, FLN and phyllochron were negatively correlated, FLN was higher in the warmer year and positively correlated with time to anthesis while phyllochron was not affected by temperature and had no correlation with time to anthesis. Under longer photoperiod, phyllochron was higher in the warmer year and time to anthesis was positively correlated with both FLN and phyllochron. Conclusions. Cultivar basal thermal requirements and PR were consistent under the different studied conditions. Changes in temperature and photoperiod affected the relationship between time to anthesis, FLN and phyllochron suggesting that, although the three traits are arithmetically related, environmental conditions affect their balance.

This study aims to investigate changes in protein secondary structures (-helix, -sheet, random coils, and -turn) and macronutrient distribution in different cool-season oat varieties and steam-pressure toasting durations using synchrotron-based mid-infrared (Mid-IR) spectroscopy and Fourier Transform Infrared spectroscopy (FTIR) imaging. All oat samples, provided by the Crop Development Center at the University of Saskatchewan, were harvested over three consecutive years (2018, 2019, and 2020). The first experiment compared four oat varieties (CDC Arborg, CDC Nasser, CDC Haymaker, and Summit), while the second examined CDC Nasser oats subjected to steam-pressure toasting (SPT) at 121 for 0, 30, 60, 90, and 120 minutes. FTIR chemical imaging revealed that carbohydrates, proteins and lipids in the four oat varieties were mainly concentrated in the endosperm, aleurone layer and embryo, crease region, and remained unchanged after SPT. Peak-fitting deconvolution of the Amide I band (1700-1600 cm-1) and subsequent quantitative analysis revealed that the four oat varieties exhibited broadly similar protein secondary structure profiles, with statistically significant but subtle variety effects detected for -helix (P = 0.026), -turn (P = 0.047), and the -helix to -sheet ratio (P = 0.048), however, -sheet and random coil proportions did not differ significantly among varieties. In contrast, SPT induced pronounced structural rearrangements, with significant increase in -sheet proportion (P = 0.003) and significant decreases in random coil content (P = 0.026). Notably, 30 minutes of toasting was sufficient to significantly increase -sheet and decrease the random coil contents. These changes are consistent with heat-induced protein denaturation and intermolecular -sheet aggregation, where thermal energy breaks the hydrogen bonds that stabilize the disordered random coil conformation, causing the unfolded polypeptide chains to reassemble into highly ordered -sheet aggregates. After SPT, the peak centers of Amide I and II bands shifted to lower wavenumbers and both bands broadened while their intensities were maintained, reflecting the reorganization of the remaining protein into -sheet aggregates rather than any loss of amide-active protein. These findings suggest that, although genotype has a relatively minor effect on the protein secondary structure of oats, hydrothermal treatments fundamentally reorganize the protein matrix from a disordered to an ordered conformation, which may have implications for protein digestibility, solubility, and nutritional function.

Ultraviolet-C (UV-C) radiation can be highly damaging to plants, yet its effects on gibberellin (GA) homeostasis are not well understood. In this study, we show that short daily UV-C pulse treatments (12 s, 1,200 J m-2) applied for seven days reduce plant height and delay flowering in Arabidopsis thaliana. Endogenous levels of the GA biosynthesis precursors GA12, GA53, GA15, and GA24, the bioactive GA4, and the GA catabolites GA34 and GA110 are all lower in UV-C treated plants than in untreated controls. These changes were accompanied by lower transcript levels of the GA biosynthesis genes KS, GA13ox1, GA20ox1, and GA3ox1, together with opposing changes in the expression of GA2ox genes. Exogenous GA4 restores growth in UV-C-treated plants, suggesting that reduced GA availability contributes to UV-C-induced growth inhibition. Consistent with this finding, the GA-signalling mutant gdella and the GA-biosynthesis mutants kao1 and kao2 show strongly reduced UV-C responses. Together, these findings highlight the importance of GA metabolism and signalling in the developmental response to repeated UV-C exposure, and suggest that exposure regimen influences the dynamics of UV-C-induced hormonal responses.

Although the biosynthetic pathways of benzylisoquinoline alkaloids (BIAs) have been extensively investigated in several plant species, their transcriptional regulatory mechanisms remain only partially understood. Jasmonate (JA)-responsive group IX APETALA2/Ethylene Responsive Factor (AP2/ERF) transcription factors (TFs) are well-known regulators of specialized plant metabolism, including the biosynthesis of various alkaloids. However, their specific roles in BIA biosynthesis remain largely elusive. Here, we isolated five novel group IX AP2/ERF TFs, designated Benzylisoquinoline alkaloid Jasmonate-responsive AP2/ERF (BJE1-5), from Coptis japonica. Phylogenetic analysis revealed that Benzylisoquinoline alkaloid Jasmonate-responsive AP2/ERF (BJE) proteins belong to subclades distinct from group IXa, which contains well-known AP2/ERF TFs involved in alkaloid biosynthesis. Transient expression analyses in C. japonica protoplasts demonstrated that certain BJEs, particularly CjBJE3 and CjBJE5, positively regulated BIA biosynthetic genes through a mutual regulatory network among BJE members. Moreover, CjBJE3 expression was regulated by CjbHLH1, a unique-type basic helix-loop-helix (bHLH) TF specific to BIA-producing plants. Furthermore, heterologous expression of CjBJE3 and CjBJE5 in cultured Eschscholzia californica cells significantly enhanced the overall BIA production, particularly by increasing end-product benzophenanthridine BIAs, highlighting several uncharacterized biosynthetic genes clustered in the genome. Our findings suggest that BIA-producing species have developed a specific regulatory network comprised of CjbHLH1 and BJE TFs, providing valuable clues for identifying novel biosynthetic enzymes.

The rejection of self-incompatible (SI) Brassica pollen is mediated by three signaling branches that function in parallel in the stigma. The recognition of SI pollen by the stigma S-Receptor Kinase (SRK) results in activation of the ARM-Repeat-Containing 1 E3 ubiquitin ligase (ARC1) which mediates the degradation of compatibility factors, the FERONIA (FER) and ANJEA (ANJ) receptor kinases that induces ROS accumulation to inhibitory levels and the M Locus Protein Kinase (MLPK) which may also be connected to ROS production. Arabidopsis self-incompatibility is regulated by SRK as well, but the signaling events downstream of SRK following SI pollen perception are less well-understood. In this study, we evaluated the requirements of FER, ANJ and HERCULES RECEPTOR KINASE 1 (HERK1) for SI pollen rejection in the transgenic Arabidopsis thaliana SI-Col-0{psi} srka-1 line. The{psi} srka-1 T-DNA disrupting the expression of the endogenous{psi} SRKA gene was crossed into SI-Col-0 to prevent any potential SRK transgene silencing. T-DNA mutants for FER and ANJ/HERK1 were then crossed into the SI-Col-0{psi} srka-1 line. Using standard assays for pollen-stigma interactions, the SI phenotypes were assessed for the SI-Col-0 fer, SI-Col-0 anj-1 and SI-Col-0 anj-1 herk1-1 lines. Our results presented here indicated that FER and ANJ are not required in the stigma for Arabidopsis SI pollen rejection, further providing evidence for a divergence in the SI downstream signaling pathway in Arabidopsis.

GOLDEN2-like (GLK) transcription factors are key regulators of chloroplast differentiation and photosynthetic gene expression. The classical yellow mutation in Japanese morning glory (Ipomoea nil) produces yellowish-green leaves, whereas an unstable allele, yellow-mutable, produces green somatic sectors on a yellowish-green background. The gene responsible for these mutations was identified as InGLK, which encodes a GOLDEN2-like transcription factor. The stable yellow mutant carried a 4-bp frameshift insertion in InGLK, whereas two yellow-mutable lines carried the Tpn1-family transposon Tpn12 in intron 5. Excision of Tpn12 in germinal revertants left short footprints and restored the green leaf phenotype. Genome searches identified InGLK as the sole GLK gene in I. nil. Pigment analysis of green somatic reversion sectors and yellowish-green background areas showed that most of the measured photosynthetic pigments were significantly reduced in the yellowish-green background, whereas the chlorophyll a/b ratio was unchanged. Chloroplasts in the yellowish-green tissue retained thylakoid-like membranes and starch granule-like structures but had less distinct grana-like stacks and sparse stromal lamellae-like structures. Wild-type-like chloroplast ultrastructure was restored in germinal revertants. These findings show that loss of function of a single-copy GLK gene broadly reduces photosynthetic pigment accumulation and alters chloroplast internal membrane organization. The yellow mutants of I. nil therefore provide a genetic system for examining non-redundant GLK function.

O_LIThis study investigated whether PEN2/BGLU26 has uniquely evolved as an indole glucosinolate-hydrolysing myrosinase required for Arabidopsis thaliana pre-invasive immunity, or whether related myrosinases can replace its function when targeted to the same subcellular context. C_LIO_LIPEN2-homologous and other selected myrosinases from A. thaliana and Brassica rapa were expressed in the pen2-2 mutant background using a PEN2-like targeting strategy. The resulting lines were assessed by gene expression, protein accumulation, metabolite analysis and pathogen resistance assays. In parallel, targeted mutagenesis, structural comparison and phylogenetic analysis were used to examine molecular and evolutionary features of PEN2-related myrosinases. C_LIO_LIAtBGLU27 and BrBABG.a, but not AtBGLU18, AtBGLU23 or AtBGLU28, partially restored indole glucosinolate hydrolysis and resistance to Colletotrichum tropicale in pen2-2. Unlike AtPEN2, both enzymes acted mainly constitutively and showed distinct substrate preferences. PEN2, BGLU27 and BABG proteins lacked conserved post-translational modification sites, including residues associated with a conserved disulfide bond. Restoring this disulfide bond in AtPEN2 abolished its activity. C_LIO_LIPEN2-related myrosinases form an evolutionarily distinct BGLU lineage associated with indole glucosinolate metabolism in Brassicales. Loss of the conserved disulfide bond appears to be required for PEN2 activity, whereas additional PEN2-specific regulatory features are needed for pathogen-triggered, rather than constitutive, glucosinolate metabolism. C_LI