Field and genetic evidence support the photosynthetic performance index (PIABS) as an indicator of rice grain yield.

The rice breeding process for grain yield could be effectively enhanced by developing efficient tools that accelerate plant selection through the rapid determination of reliable predictors. In this study, we have described various associations between grain yield and photosynthetic parameters, which can be easily and quickly obtained using a non-invasive technique on the flag leaf during the anthesis stage. Among the analyzed photosynthetic parameters, the photosynthetic performance index (PIABS) stood out due to its strong association with grain yield. A genome-wide association analysis conducted on plants from a rice diversity panel at the tillering stage revealed the presence of a quantitative trait locus on chromosome 9. This locus was characterized by a group of candidate chloroplastic genes that exhibited contrasting haplotypes for PIABS. An analysis of these haplotypes revealed a clear division into two groups. One group consisted of haplotypes linked to high values of PIABS, which were predominantly associated with Japonica spp. subpopulations. The other group consisted of haplotypes linked to low values of PIABS, which were exclusively associated with Indica spp. subpopulations. Japonica spp. genotypes exhibited higher values in the yield component panicle weight compared with the Indica spp. genotypes. The findings of this study indicate that PIABS could serve as an early predictor of yield parameters during the tillering stage in rice breeding processes.

Patterns of physiological parameters and nitrogen partitioning in flag leaf explain differential grain protein content in rice.

The grain protein content (GPC) in rice is low, and more efforts with agronomic and molecular approaches were performed to increase them. However, the rice research focusing on the plant physiological behaviour that modulates the phenomenon of grain protein filling is very scarce. This work contains physiological parameters related to photosynthetic activity in the flag leaf in the grain filling period and N partitioning assays of high (Nutriar) and traditional (Camba) GPC cultivars. Results indicated a higher photosynthetic capacity, a better capacity to provide CO2 to the chloroplast and a healthier PSII structure in Camba relative to Nutriar. Chlorophyll fluorescence parameters decreased more steeply over time in the high protein variety, and a strong negative correlation was observed between GPC and PSII structure parameters. N content in the flag leaf at anthesis showed lower values and higher remobilisation during the grain filling period in Nutriar compared to Camba. The results of this work suggested that the inactivation of some PSII structures in higher GPC cultivars is associated with N remobilisation and would contribute to an increase in the free N available to be translocated to the grain.

Stem canker caused by Phomopsis spp. Induces changes in polyamine levels and chlorophyll fluorescence parameters in pecan leaves.

Pecan plants are attacked by the fungus Phomopsis spp. that causes stem canker, a serious and emerging disease in commercial orchards. Stem canker, which has been reported in several countries, negatively affects tree canopy health, eventually leading to production losses. The purpose of this study was to inquire into the physiology of pecan plants under stem canker attack by Phomopsis spp. To this end, pecan plants were inoculated with an isolate of Phomopsis spp. and several parameters, such as polyamines, proline, sugars, starch, chlorophyll fluorescence and canopy temperature were analysed. Under artificial inoculation, a high disease incidence was observed with symptoms similar to those in plants showing stem canker under field conditions. Furthermore, the infected stem showed dead tissue with brown necrotic discolouration in the xylem tissue. The free polyamines putrescine, spermidine, and spermine were detected and their levels decreased as leaves aged in the infected plants with respect to the controls. Chlorophyll fluorescence parameters, such as Sm, ψEO, and QbRC decreased under plant infection and therefore the K-band increased. Canopy temperature and proline content increased in the infected plants with respect to the controls while sugar content decreased. These data suggest that stem canker caused by Phomopsis spp. induces physiological changes that are similar to those observed in plants under drought stress. To our knowledge, this is the first study that documents the physiological and biochemical effects derived from pecan-Phomopsis interaction.

Close relationship between the state of the oxygen evolving complex and rice cold stress tolerance.

The results of the present work suggested a relationship between the growth stability and functional/structural parameters associated to the primary photochemistry and oxygen evolving complex (OEC) in tolerant rice plants under suboptimal low temperatures (SLT) stress. This was concluded from the absence of changes in net photosynthetic rate and in fraction of reaction centers to reduce quinone A, and very small changes in P680 efficiency to trap and donate electrons to quinone A and in fraction of active OEC in tolerant plants under cold stress but not in sensitive plants. The SLT stress also induced OEC activity limitations in both genotypes, but in a greater extent in sensitive plants. However, an assay using an artificial electron donor to replace OEC indicated that the P680+ capacity to accept electrons was not altered in both genotypes under SLT stress from the beginning of the stress treatment, suggesting that the OEC structure stability is related to rice SLT tolerance to sustain the photosynthesis. This hypothesis was also supported by the fact that tolerant plants but not sensitive plants did not alter the gene expression and protein content of PsbP under SLT stress, an OEC subunit with a role in stabilizing of OEC structure.

Differential effects of azinphos-methyl and chlorpyrifos on polyamine oxidative metabolism during the embryonic development of Rhinella arenarum and its relation to oxidative stress.

The organophosphorus pesticides azinphos-methyl (AZM) and chlorpyrifos (CPF) exert their toxic action by inhibition of acetylcholinesterase, but non-target processes such as polyamine metabolism can also be affected. Our objective was to evaluate the effects of different concentrations of AZM (0.5-, 2- and 9 mg L-1) and CPF (0.5- and 1 mg L-1) on polyamine oxidative metabolism along Rhinella arenarum embryonic development and to explore its relationship to oxidative stress. Free and conjugated polyamines were measured by HPLC. The activity of spermine oxidase (SMOX), N1-acetylpolyamine oxidase (PAOX) and diamine oxidase (DAO) were measured through kinetic spectrofluorometry. Free putrescine and spermine were significantly increased in open mouth embryos exposed to AZM. Free polyamine levels were not affected by CPF exposure. In embryos exposed to AZM, DAO was increased in tail bud stage and SMOX was increased in open mouth stage, while embryos exposed to CPF showed an increase of PAOX activity in tail bud stage and a decrease of DAO and SMOX activity in open mouth stage. Polyamine levels and oxidative degradation enzymes respond differently if R. arenarum embryos are exposed to AZM or CPF, despite that both insecticides belong to the same chemical family. The early increase of DAO and PAOX would play a protective role to guarantee the normal progression of embryonic development. The increased production of reactive species might contribute to an oxidative stress situation generated by exposure to the insecticides and to the alteration of the antioxidant defense system. In tail bud stage embryos, PAOX and SMOX were positively correlated to acetylcholinesterase activity and reduced glutathione levels (GSH), and negatively correlated to the antioxidant enzymes catalase (CAT) and glutathione S-transferase (GST). In complete operculum embryos, a negative correlation between antioxidant parameters and polyamine levels and polyamine oxidative metabolism was observed, except for SMOX, which showed a low positive correlation with CAT and GSH and a negative correlation to PAOX and DAO. We suggest the use of DAO and PAOX as biomarkers of exposure to AZM and CPF, respectively, as they respond earlier than the classical biomarker acetylcholinesterase.

Redox homeostasis in the growth zone of the rice leaf plays a key role in cold tolerance.

We analysed the cellular and molecular changes in the leaf growth zone of tolerant and sensitive rice varieties in response to suboptimal temperatures. Cold reduced the final leaf length by 35% and 51% in tolerant and sensitive varieties, respectively. Tolerant lines exhibited a smaller reduction of the leaf elongation rate and greater compensation by an increased duration of leaf growth. Kinematic analysis showed that cold reduced cell production in the meristem and the expansion rate in the elongation zone, but the latter was compensated for by a doubling of the duration of cell expansion. We performed iTRAQ proteome analysis on proliferating and expanding parts of the leaf growth zone. We identified 559 and 542 proteins, of which 163 and 210 were differentially expressed between zones, and 96 and 68 between treatments, in the tolerant and sensitive lines, respectively. The categories protein biosynthesis and redox homeostasis were significantly overrepresented in the up-regulated proteins. We therefore measured redox metabolites and enzyme activities in the leaf growth zone, demonstrating that tolerance of rice lines to suboptimal temperatures correlates with the ability to up-regulate enzymatic antioxidants in the meristem and non-enzymatic antioxidants in the elongation zone.

Polyamines and Legumes: Joint Stories of Stress, Nitrogen Fixation and Environment.

Polyamines (PAs) are natural aliphatic amines involved in many physiological processes in almost all living organisms, including responses to abiotic stresses and microbial interactions. On other hand, the family Leguminosae constitutes an economically and ecologically key botanical group for humans, being also regarded as the most important protein source for livestock. This review presents the profuse evidence that relates changes in PAs levels during responses to biotic and abiotic stresses in model and cultivable species within Leguminosae and examines the unreviewed information regarding their potential roles in the functioning of symbiotic interactions with nitrogen-fixing bacteria and arbuscular mycorrhizae in this family. As linking plant physiological behavior with "big data" available in "omics" is an essential step to improve our understanding of legumes responses to global change, we also examined integrative MultiOmics approaches available to decrypt the interface legumes-PAs-abiotic and biotic stress interactions. These approaches are expected to accelerate the identification of stress tolerant phenotypes and the design of new biotechnological strategies to increase their yield and adaptation to marginal environments, making better use of available plant genetic resources.

The fungal endophyte Fusarium solani provokes differential effects on the fitness of two Lotus species.

The interactions established between plants and endophytic fungi span a continuum from beneficial to pathogenic associations. The aim of this work was to isolate potentially beneficial fungal endophytes in the legume Lotus tenuis and explore the mechanisms underlying their effects. One of the nine fungal strains isolated was identified as Fusarium solani and shows the highest phosphate-solubilisation activity, and also grows endophytically in roots of L. japonicus and L. tenuis. Interestingly, fungal invasion enhances plant growth in L. japonicus but provokes a contrasting effect in L. tenuis. These differences were also evidenced when the rate of photosynthesis as well as sugars and K contents were assessed. Our results indicate that the differential responses observed are due to distinct mechanisms deployed during the establishment of the interactions that involve the regulation of photosynthesis, potassium homeostasis, and carbohydrate metabolism. These responses are employed by these plant species to maintain fitness during the endophytic interaction.

Heat treatment alleviates the growth and photosynthetic impairment of transplastomic plants expressing Leishmania infantum Hsp83-Toxoplasma gondii SAG1 fusion protein.

Previously, we showed that transplastomic tobacco plants expressing the LiHsp83-SAG1 fusion protein displayed a chlorotic phenotype and growth retardation, while plants expressing the SAG1 and GRA4 antigens alone did not. We conducted a comprehensive examination of the metabolic and photosynthetic parameters that could be affecting the normal growth of LiHsp83-SAG1 plants in order to understand the origin of these pleiotropic effects. These plants presented all photosynthetic pigments and parameters related to PSII efficiency significantly diminished. However, the expression of CHLI, RSSU and LHCa/b genes did not show significant differences between LiHsp83-SAG1 and control plants. Total protein, starch, and soluble sugar contents were also greatly reduced in LiHsp83-SAG1 plants. Since Hsp90 s are constitutively expressed at much higher concentrations at high temperatures, we tested if the fitness of LiHsp83-SAG1 over-expressing LiHsp83 would improve after heat treatment. LiHsp83-SAG1 plants showed an important alleviation of their phenotype and an evident recovery of the PSII function. As far as we know, this is the first report where it is demonstrated that a transplastomic line performs much better at higher temperatures. Finally, we detected that LiHsp83-SAG1 protein could be binding to key photosynthesis-related proteins at 37 °C. Our results suggest that the excess of this molecular chaperone could benefit the plant in a possible heat shock and prevent the expected denaturation of proteins. However, the LiHsp83-SAG1 protein content was weakly decreased in heat-treated plants. Therefore, we cannot rule out that the alleviation observed at 37 °C may be partially due to a reduction of the levels of the recombinant protein.

Modulation of plant and bacterial polyamine metabolism during the compatible interaction between tomato and Pseudomonas syringae.

The polyamines putrescine, spermidine and spermine participate in a variety of cellular processes in all organisms. Many studies have shown that these polycations are important for plant immunity, as well as for the virulence of diverse fungal phytopathogens. However, the polyamines' roles in the pathogenesis of phytopathogenic bacteria have not been thoroughly elucidated to date. To obtain more information on this topic, we assessed the changes in polyamine homeostasis during the infection of tomato plants by Pseudomonas syringae. Our results showed that polyamine biosynthesis and catabolism are activated in both tomato and bacteria during the pathogenic interaction. This activation results in the accumulation of putrescine in whole leaf tissues, as well as in the apoplastic fluids, which is explained by the induction of its synthesis in plant cells and also on the basis of its excretion by bacteria. We showed that the excretion of this polyamine by P. syringae is stimulated under virulence-inducing conditions, suggesting that it plays a role in plant colonization. However, no activation of bacterial virulence traits or induction of plant invasion was observed after the exogenous addition of putrescine. In addition, no connection was found between this polyamine and plant defence responses. Although further research is warranted to unravel the biological functions of these molecules during plant-bacterial interactions, this study contributes to a better understanding of the changes associated with the homeostasis of polyamines during plant pathogenesis.

Rice tolerance to suboptimal low temperatures relies on the maintenance of the photosynthetic capacity.

The purpose of this research was to identify differences between two contrasting rice cultivars in their response to suboptimal low temperatures stress. A transcriptomic analysis of the seedlings was performed and results were complemented with biochemical and physiological analyses. The microarray analysis showed downregulation of many genes related with PSII and particularly with the oxygen evolving complex in the sensitive cultivar IR50. Complementary studies indicated that the PSII performance, the degree of oxygen evolving complex coupling with the PSII core and net photosynthetic rate diminished in this cultivar in response to the stress. However, the tolerant cultivar Koshihikari was able to maintain its energy equilibrium by sustaining the photosynthetic capacity. The increase of oleic acid in Koshihikari could be related with membrane remodelling of the chloroplasts and hence contribute to tolerance. Overall, these results work as a ground for future analyses that look forward to characterize possible mechanisms to tolerate this stress.

Polyamine Metabolism Responses to Biotic and Abiotic Stress.

Plants have developed different strategies to cope with the environmental stresses they face during their life cycle. The responses triggered under these conditions are usually characterized by significant modifications in the metabolism of polyamines such as putrescine, spermidine, and spermine. Several works have demonstrated that a fine-tuned regulation of the enzymes involved in the biosynthesis and catabolism of polyamines leads to the increment in the concentration of these compounds. Polyamines exert different effects that could help plants to deal with stressful conditions. For instance, they interact with negatively charged macromolecules and regulate their functions, they may act as compatible osmolytes, or present antimicrobial activity against plant pathogens. In addition, they have also been proven to act as regulators of gene expression during the elicitation of stress responses. In this chapter, we reviewed the information available till date in relation to the roles played by polyamines in the responses of plants during biotic and abiotic stress.

Reactive oxygen species generated in chloroplasts contribute to tobacco leaf infection by the necrotrophic fungus Botrytis cinerea.

Reactive oxygen species (ROS) play fundamental roles in plant responses to pathogen infection, including modulation of cell death processes and defense-related gene expression. Cell death triggered as part of the hypersensitive response enhances resistance to biotrophic pathogens, but favors the virulence of necrotrophs. Even though the involvement of ROS in the orchestration of defense responses is well established, the relative contribution of specific subcellular ROS sources to plant resistance against microorganisms with different pathogenesis strategies is not completely known. The aim of this work was to investigate the role of chloroplastic ROS in plant defense against a typical necrotrophic fungus, Botrytis cinerea. For this purpose, we used transgenic Nicotiana tabacum (tobacco) lines expressing a plastid-targeted cyanobacterial flavodoxin (pfld lines), which accumulate lower chloroplastic ROS in response to different stresses. Tissue damage and fungal growth were significantly reduced in infected leaves of pfld plants, as compared with infected wild-type (WT) counterparts. ROS build-up triggered by Botrytis infection and associated with chloroplasts was significantly decreased (70-80%) in pfld leaves relative to the wild type. Phytoalexin accumulation and expression of pathogenesis-related genes were induced to a lower degree in pfld plants than in WT siblings. The impact of fungal infection on photosynthetic activity was also lower in pfld leaves. The results indicate that chloroplast-generated ROS play a major role in lesion development during Botrytis infection. This work demonstrates that the modulation of chloroplastic ROS levels by the expression of a heterologous antioxidant protein can provide a significant degree of protection against a canonical necrotrophic fungus.

The alkaline tolerance in Lotus japonicus is associated with mechanisms of iron acquisition and modification of the architectural pattern of the root.

The response of fifty-four Lotus japonicus ecotypes, and of six selected ecotypes was investigated under alkaline conditions. Sensitive, but not tolerant ecotypes, showed interveinal chlorosis under all alkalinity conditions and high mortality under extreme alkalinity. Interveinal chlorosis was associated with Fe deficiency, as a reduced Fe2+ shoot content was observed in all sensitive ecotypes. In addition, some showed a decline in photosynthesis rate and PSII performance compared to the control. In contrast, some tolerant ecotypes did not change these parameters between treatments. Alkaline tolerance could be explained by a mechanism of Fe acquisition and a root structural modification. This conclusion was based on the fact that all tolerant, but not the sensitive ecotypes, presented high ferric reductase oxidase activity under alkaline stress compared to the control, and a Herringbone root pattern modification. On this basis, the analysis of these mechanisms of alkaline tolerance could be used in screening programs for the selection of new tolerant genotypes in the Lotus genus.

Photosynthetic responses mediate the adaptation of two Lotus japonicus ecotypes to low temperature.

Lotus species are important forage legumes due to their high nutritional value and adaptability to marginal conditions. However, the dry matter production and regrowth rate of cultivable Lotus spp. is drastically reduced during colder seasons. In this work, we evaluated the chilling response of Lotus japonicus ecotypes MG-1 and MG-20. No significant increases were observed in reactive oxygen species and nitric oxide production or in lipid peroxidation, although a chilling-induced redox imbalance was suggested through NADPH/NADP(+) ratio alterations. Antioxidant enzyme catalase, ascorbate peroxidase, and superoxide dismutase activities were also measured. Superoxide dismutase, in particular the chloroplastic isoform, showed different activity for different ecotypes and treatments. Stress-induced photoinhibition also differentially influenced both ecotypes, with MG-1 more affected than MG-20. Data showed that the D2 PSII subunit was more affected than D1 after 1 d of low temperature exposure, although its protein levels recovered over the course of the experiment. Interestingly, D2 recovery was accompanied by improvements in photosynthetic parameters (Asat and Fv/Fm) and the NADPH/NADP(+) ratio. Our results suggest that the D2 protein is involved in the acclimation response of L. japonicus to low temperature. This may provide a deeper insight into the chilling tolerance mechanisms of the Lotus genus.

Transcriptome Response Mediated by Cold Stress in Lotus japonicus.

Members of the Lotus genus are important as agricultural forage sources under marginal environmental conditions given their high nutritional value and tolerance of various abiotic stresses. However, their dry matter production is drastically reduced in cooler seasons, while their response to such conditions is not well studied. This paper analyzes cold acclimation of the genus by studying Lotus japonicus over a stress period of 24 h. High-throughput RNA sequencing was used to identify and classify 1077 differentially expressed genes, of which 713 were up-regulated and 364 were down-regulated. Up-regulated genes were principally related to lipid, cell wall, phenylpropanoid, sugar, and proline regulation, while down-regulated genes affected the photosynthetic process and chloroplast development. Together, a total of 41 cold-inducible transcription factors were identified, including members of the AP2/ERF, NAC, MYB, and WRKY families; two of them were described as putative novel transcription factors. Finally, DREB1/CBFs were described with respect to their cold stress expression profiles. This is the first transcriptome profiling of the model legume L. japonicus under cold stress. Data obtained may be useful in identifying candidate genes for breeding modified species of forage legumes that more readily acclimate to low temperatures.

Transcriptional regulation of 9-cis-epoxycarotenoid dioxygenase (NCED) gene by putrescine accumulation positively modulates ABA synthesis and drought tolerance in Lotus tenuis plants.

The accumulation of putrescine (Put) and increased arginine decarboxylase (ADC, EC 4.1.1.19) activity levels in response to osmotic stress has been reported; however, the biological meaning of this increase remains unclear. To obtain new insights into these questions, we studied the drought response of a transgenic Lotus tenuis line that expresses the oat ADC gene, which is driven by the stress-inducible pRD29A promoter. This line contains high levels of Put with no changes in spermidine and spermine contents, even under water deficits. Our results indicate that the biochemical and morphological responses to dehydration correlate with the Put level and provide evidence that Put controls the ABA content in response to drought by modulating ABA biosynthesis at the transcriptional level.

A succinate dehydrogenase flavoprotein subunit-like transcript is upregulated in Ilex paraguariensis leaves in response to water deficit and abscisic acid.

Ilex paraguariensis plants were subjected to progressive soil water deficit, and differential display (DD) was used to analyse gene expression in leaves to characterise physiological responses to mild and severe water deficits. A cDNA fragment showing strong homology with the flavoprotein subunit (SDH1) of succinate:ubiquinone oxidoreductase (succinate dehydrogenase, SDH, EC 1.3.5.1) was upregulated in plants exposed to drought. Quantitative real-time PCR revealed that the SDH1-like transcript level began to increase when the leaf relative water content (RWC) decreased to 78% and peaked when the RWC dropped to 57%. A correlation between abscisic acid (ABA) concentration and variations in transcript levels was assessed by GC-SIM. After rehydration, SDH1 mRNA and ABA returned to their initial levels. In stressed leaves sprayed with ABA SDH1 mRNA accumulated in greater levels compared to stressed leaves that did not receive ABA. Moreover, the enzymatic activity of succinate dehydrogenase increased 1.5-fold in the mature leaves of ABA-treated plants. This physiological response may be related to the tendency of this species to minimise water losses through stomatal closure in the early stages of dehydration to avoid tissue desiccation. As the leaf water potential diminished due to an increase in water restriction, I. paraguariensis leaf tissues reacted by making osmotic adjustments to sustain tissue metabolic activity, which enables the recovery of photosynthesis upon re-watering. These results provide new insights concerning the linkage between plant respiration and photosynthetic metabolism that could be potentially further used in breeding programs aiming water tolerant genotypes.

Polyamine catabolism is involved in response to salt stress in soybean hypocotyls.

The possible relationship between polyamine catabolism mediated by copper-containing amine oxidase and the elongation of soybean hypocotyls from plants exposed to NaCl has been studied. Salt treatment reduced values of all hypocotyl growth parameters. In vitro, copper-containing amine oxidase activity was up to 77-fold higher than that of polyamine oxidase. This enzyme preferred cadaverine over putrescine and it was active even under the saline condition. On the other hand, saline stress increased spermine and cadaverine levels, and the in vivo copper-containing amine oxidase activity in the elongation zone of hypocotyls. The last effect was negatively modulated by the addition of the copper-containing amine oxidase inhibitor N,N'-diaminoguanidine. In turn, plants treated with the inhibitor showed a significant reduction of reactive oxygen species in the elongation zone, even in the saline situation. In addition, plants grown in cadaverine-amended culture medium showed increased hypocotyl length either in saline or control conditions and this effect was also abolished by N,N'-diaminoguanidine. Taken together, our results suggest that the activity of the copper-containing amine oxidase may be partially contributing to hypocotyl growth under saline stress, through the production of hydrogen peroxide by polyamine catabolism and reinforce the importance of polyamine catabolism and hydrogen peroxide production in the induction of salt tolerance in plants.

Polyamine oxidase activity contributes to sustain maize leaf elongation under saline stress.

The possible involvement of apoplastic reactive oxygen species produced by the oxidation of free polyamines in the leaf growth of salinized maize has been studied here. Salt treatment increased the apoplastic spermine and spermidine levels, mainly in the leaf blade elongation zone. The total activity of polyamine oxidase was up to 20-fold higher than that of the copper-containing amine oxidase. Measurements of H(2)O(2), *O(2)(-), and HO* production in the presence or absence of the polyamine oxidase inhibitors 1,19-bis-(ethylamine)-5,10,15 triazanonadecane and 1,8-diamino-octane suggest that, in salinized plants, the oxidation of free apoplastic polyamines by polyamine oxidase by would be the main source of reactive oxygen species in the elongation zone of maize leaf blades. This effect is probably due to increased substrate availability. Incubation with 200 microM spermine doubled segment elongation, whereas the addition of 1,19-bis-(ethylamine)-5,10,15 triazanonadecane and 1,8-diamino-octane to 200 microM spermine attenuated and reversed the last effect, respectively. Similarly, the addition of MnCl(2) (an *O(2)(-) dismutating agent) or the HO* scavenger sodium benzoate along with spermine, annulled the elongating effect of the polyamine on the salinized segments. As a whole, the results obtained here demonstrated that, under salinity, polyamine oxidase activity provides a significant production of reactive oxygen species in the apoplast which contributes to 25-30% of the maize leaf blade elongation.