Arbuscular mycorrhizal fungi offset NH3 emissions in temperate meadow soil under simulated warming and nitrogen deposition.

Most soil ammonia (NH3) emissions originate from soil nitrogen (N) that has been in the form of exchangeable ammonium. Emitted NH3 not only induces nutrient loss but also has adverse effects on the cycling of N and accelerates global warming. There is evidence that arbuscular mycorrhizal (AM) fungi can alleviate N loss by reducing N2O emissions in N-limited ecosystems, however, some studies have also found that global changes, such as warming and N deposition, can affect the growth and development of AM fungi and alter their functionality. Up to now, the impact of AM fungi on NH3 emissions, and whether global changes reduce the AM fungi's contribution to NH3 emissions reduction, has remained unclear. In this study, we examined how warming, N addition, and AM fungi alter NH3 emissions from high pH saline soils typical of a temperate meadow through a controlled microscopic experiment. The results showed that warming significantly increased soil NH3 emissions, but N addition and combined warming plus N addition had no impact. Inoculations with AM fungi strongly reduced NH3 emissions both under warming and N addition, but AM fungi effects were more pronounced under warming than following N addition. Inoculation with AM fungi reduced soil NH4+-N content and soil pH, and increased plant N content and soil net N mineralization rate while increasing the abundance of ammonia-oxidizing bacterial (AOB) gene. Structural equation modeling (SEM) shows that the regulation of NH3 emissions by AM fungi may be related to soil NH4+-N content and soil pH. These findings highlight that AM fungi can reduce N loss in the form of NH3 by increasing N turnover and uptake under global changes; thus, AM fungi play a vital role in alleviating the aggravation of N loss caused by global changes and in mitigating environmental pollution in the future.

Integrated metabolomic and transcriptomic strategies to reveal alkali-resistance mechanisms in wild soybean during post-germination growth stage.

MAIN CONCLUSION: The keys to alkali-stress resistance of barren-tolerant wild soybean lay in enhanced reutilization of reserves in cotyledons as well as improved antioxidant protection and organic acid accumulation in young roots. Soil alkalization of farmlands is increasingly serious, adversely restricting crop growth and endangering food security. Here, based on integrated analysis of transcriptomics and metabolomics, we systematically investigated changes in cotyledon weight and young root growth in response to alkali stress in two ecotypes of wild soybean after germination to reveal alkali-resistance mechanisms in barren-tolerant wild soybean. Compared with barren-tolerant wild soybean, the dry weight of common wild soybean cotyledons under alkali stress decreased slowly and the length of young roots shortened. In barren-tolerant wild soybean, nitrogen-transport amino acids asparagine and glutamate decreased in cotyledons but increased in young roots, and nitrogen-compound transporter genes and genes involved in asparagine metabolism were significantly up-regulated in both cotyledons and young roots. Moreover, isocitric, succinic, and L-malic acids involved in the glyoxylate cycle significantly accumulated and the malate synthetase gene was up-regulated in barren-tolerant wild soybean cotyledons. In barren-tolerant wild soybean young roots, glutamate and glycine related to glutathione metabolism increased significantly and the glutathione reductase gene was up-regulated. Pyruvic acid and citric acid involved in pyruvate-citrate metabolism increased distinctly and genes encoding pyruvate decarboxylase and citrate synthetase were up-regulated. Integrated analysis showed that the keys to alkali-stress resistance of barren-tolerant wild soybean lay in enhanced protein decomposition, amino acid transport, and lipolysis in cotyledons as well as improved antioxidant protection and organic acid accumulation in young roots. This study provides new ideas for the exploitation and utilization of wild soybean resources.

Wild soybean salt tolerance metabolic model: Assessment of storage protein mobilization in cotyledons and C/N balance in the hypocotyl/root axis.

Salt stress has become one of the main factors limiting crop yield in recent years. The post-germinative growth is most sensitive to salt stress in soybean. In this study, cultivated and wild soybeans were used for an integrated metabonomics and transcriptomics analysis to determine whether wild soybean can resist salt stress by maintaining the mobilization of stored substances in cotyledons and the balance of carbon and nitrogen in the hypocotyl/root axis (HRA). Compared with wild soybean, the growth of cultivated soybean was significantly inhibited during the post-germinative growth period under salt stress. Integrating analysis found that the breakdown products of proteins, such as glutamate, glutamic acid, aspartic acid, and asparagine, increased significantly in wild soybean cotyledons. Asparagine synthase and fumarate hydratase genes and genes encoding HSP20 family proteins were specifically upregulated. In wild soybean HRA, levels of glutamic acid, aspartic acid, asparagine, citric acid, and succinic acid increased significantly, and the glutamate decarboxylase gene and the gene encoding carbonic anhydrase in nitrogen metabolism were significantly upregulated. The metabolic model indicated that wild soybean enhanced the decomposition of stored proteins and the transport of amino acids to the HRA in cotyledons and the GABA shunt to maintain carbon and nitrogen balance in the HRA to resist salt stress. This study provided a theoretical basis for cultivating salt-tolerant soybean varieties and opened opportunities for the development of sustainable agricultural practices.

Comprehensive physiological, transcriptomic, and metabolomic analysis of the key metabolic pathways in millet seedling adaptation to drought stress.

Drought is one of the leading environmental constraints that affect the growth and development of plants and, ultimately, their yield and quality. Foxtail millet (Setaria italica) is a natural stress-resistant plant and an ideal model for studying plant drought resistance. In this study, two varieties of foxtail millet with different levels of drought resistance were used as the experimental material. The soil weighing method was used to simulate drought stress, and the differences in growth, photosynthetic physiology, metabolite metabolism, and gene transcriptional expression under drought stress were compared and analyzed. We aimed to determine the physiological and key metabolic regulation pathways of the drought-tolerant millet in resistance to drought stress. The results showed that drought-tolerant millet exhibited relatively stable growth and photosynthetic parameters under drought stress while maintaining a relatively stable level of photosynthetic pigments. The metabolomic, transcriptomic, and gene co-expression network analysis confirmed that the key to adaptation to drought by millet was to enhance lignin metabolism, promote the metabolism of fatty acids to be transformed into cutin and wax, and improve ascorbic acid circulation. These findings provided new insights into the metabolic regulatory network of millet adaptation to drought stress.

Remodeling and protecting the membrane system to resist phosphorus deficiency in wild soybean (Glycine soja) seedling leaves.

MAIN CONCLUSION: The poor-soil-tolerant wild soybean resist phosphorus deficiency by remodeling membrane lipids to reuse phosphorus. The plants synthesize phenolic acids and flavonoids to remove reactive oxygen species and protect membrane stability. Poor soil largely limits plant yields, and the development and utilization of high-quality wild plant resources is an effective approach to resolving this problem. Two ecotypes of wild soybean were used as experimental materials in this experiment. We integrated metabolomics and transcriptomics to determine whether wild soybean (Glycine soja) could resist phosphorus deficiency by remodeling and protecting its membrane system. Under phosphorus-deficient conditions, the plant height and aboveground fresh and dry weight of poor-soil-tolerant wild soybean seedlings were less inhibited than those in common wild soybean. In poor-soil-tolerant wild soybean seedling leaves, the glycerol-3-phosphate content decreased significantly, while caffeic acid, ferulic acid, shikimic acid, phenylalanine, tyrosine, and tryptophan increased significantly. ß-Glucosidase and chalcone synthase genes and those that encode SQD2, a crucial enzyme in thiolipid biosynthesis, were specifically up-regulated, whereas the glucosyltransferase UGT74B1 gene was down-regulated. The poor-soil-tolerant wild soybean enhanced glycerolipid metabolism to decompose phospholipids and release phosphorus for reuse to improve resistance to phosphorus deficiency. The plants synthesized thiolipids to replace phospholipids and maintain membrane structure integrity and inhibited glucosinolate biosynthesis to promote phenylpropanoid biosynthesis, leading to the production of phenolic acids and flavonoids that removed reactive oxygen species and protected membrane system stability. The experiments evaluated and provided insight into the innovative utilization of wild soybean germplasm resources.

Suppression of Arbuscular Mycorrhizal Fungi Aggravates the Negative Interactive Effects of Warming and Nitrogen Addition on Soil Bacterial and Fungal Diversity and Community Composition.

We examined the impacts of warming, nitrogen (N) addition, and suppression of arbuscular mycorrhizal fungi (AMF) on soil bacterial and fungal richness and community composition in a field experiment. AMF root colonization and the concentration of an AMF-specific phospholipid fatty acid (PLFA) were significantly reduced after the application of the fungicide benomyl as a soil drench. Warming and N addition had no independent effects but interactively decreased soil fungal richness, while warming, N addition, and AMF suppression together reduced soil bacterial richness. Soil bacterial and fungal species diversity was lower with AMF suppression, indicating that AMF suppression has a negative effect on microbial diversity. Warming and N addition decreased the net loss of plant species and the plant species richness, respectively. AMF suppression reduced plant species richness and the net gain of plant species but enhanced the net loss of plant species. Structural equation modeling (SEM) demonstrated that the soil bacterial community responded to the increased soil temperature (ST) induced by warming and the increased soil available N (AN) induced by N addition through changes in AMF colonization and plant species richness; ST directly affected the bacterial community, but AN affected both the soil bacterial and fungal communities via AMF colonization. In addition, higher mycorrhizal colonization increased the plant species richness by increasing the net gains in plant species under warming and N addition. IMPORTANCE AMF can influence the composition and diversity of plant communities. Previous studies have shown that climate warming and N deposition reduce the effectiveness of AMF. However, how AMF affect soil bacterial and fungal communities under these global change drivers is still poorly understood. A 4-year field study revealed that AMF suppression decreased bacterial and fungal diversity irrespective of warming or N addition, while AMF suppression interacted with warming or N addition to reduce bacterial and fungal richness. In addition, bacterial and fungal community compositions were determined by mycorrhizal colonization, which was regulated by soil AN and ST. These results suggest that AMF suppression can aggravate the severe losses to native soil microbial diversity and functioning caused by global changes; thus, AMF play a vital role in maintaining belowground ecosystem stability in the future.

Physiological characteristics and metabolomics reveal the tolerance mechanism to low nitrogen in Glycine soja leaves.

To explore the regulatory mechanisms involved in the adaption to nitrogen (N) deficiency of wild soybean, the ion balance, photosynthetic characteristics, metabolic and transcriptional changes in leaves of common and low N (LN)-tolerant wild soybean seedlings under LN stress were determined. The LN-tolerant wild soybean seedlings showed a stronger ability to maintain photosynthesis and nutrient balance than common wild soybean. A total of 52 differentially accumulated metabolites, mainly related to carbon and N metabolism, were identified between the control and the LN treatment group. In general, tricarboxylic acid (TCA) cycle, shikimic acid pathway, synthetase/glutamate synthase (GS/GOGAT) cycle and accumulation of most organic acids were enhanced in LN-tolerant wild soybean, while reduced in common wild soybean under LN stress compared with their respective control group. Moreover, glycolysis, sugar and polyol and fatty acid metabolism increased in both wild soybean genotypes, and increased more in LN-tolerant wild soybean. A total of 3381 differentially expressed genes (DEGs) were identified in leaves of both wild soybean genotypes and the expressed level of DEGs associated with sugars, polyols, fatty acids and energy metabolism was significantly higher in LN-tolerant wild soybean than in common wild soybean, consistent with changes in metabolite level. Our results suggest new ideas for the study of LN tolerance of wild soybean and provide a theoretical basis for development and utilization of wild soybean resources.

The subcellular localization of two isopentenyl diphosphate isomerases in rice suggests a role for the endoplasmic reticulum in isoprenoid biosynthesis.

KEY MESSAGE: Both OsIPPI1 and OsIPPI2 enzymes are found in the endoplasmic reticulum, providing novel important insights into the role of this compartment in the synthesis of MVA pathway isoprenoids. Isoprenoids are synthesized from the precursor's isopentenyl diphosphate (IPP) and dimethylallyl diphosphosphate (DMAPP), which are interconverted by the enzyme isopentenyl diphosphate isomerase (IPPI). Many plants express multiple isoforms of IPPI, the only enzyme shared by the mevalonate (MVA) and non-mevalonate (MEP) pathways, but little is known about their specific roles. Rice (Oryza sativa) has two IPPI isoforms (OsIPPI1 and OsIPPI2). We, therefore, carried out a comprehensive comparison of IPPI gene expression, protein localization, and isoprenoid biosynthesis in this species. We found that OsIPPI1 mRNA was more abundant than OsIPPI2 mRNA in all tissues, and its expression in de-etiolated leaves mirrored the accumulation of phytosterols, suggesting a key role in the synthesis of MVA pathway isoprenoids. We investigated the subcellular localization of both isoforms by constitutively expressing them as fusions with synthetic green fluorescent protein. Both proteins localized to the endoplasmic reticulum (ER) as well as peroxisomes and mitochondria, whereas only OsIPPI2 was detected in plastids, due to an N-terminal transit peptide which is not present in OsIPPI1. Despite the plastidial location of OsIPPI2, the expression of OsIPPI2 mRNA did not mirror the accumulation of chlorophylls or carotenoids, indicating that OsIPPI2 may be a redundant component of the MEP pathway. The detection of both OsIPPI isoforms in the ER indicates that DMAPP can be synthesized de novo in this compartment. Our work shows that the ER plays an as yet unknown role in the synthesis of MVA-derived isoprenoids, with important implications for the metabolic engineering of isoprenoid biosynthesis in higher plants.

Physiological and metabolomics analyses of young and old leaves from wild and cultivated soybean seedlings under low-nitrogen conditions.

BACKGROUND: It is critical to study the low nitrogen tolerance in wild soybean with extensive genetic diversity for improving cultivated soybean nitrogen use efficiency. Focusing on plant young and old leaves could provide new insights to low nitrogen tolerance research. This study compared the low nitrogen group with the control group on physiological and metabolomics changes in young and old leaves, respectively, then analyzed and compared the differences of these changes between cultivated and wild soybean. This study aimed to provide a theoretical basis for the molecular mechanism of soybean low nitrogen stress tolerance. RESULTS: Wild soybean was less affected by low-nitrogen stress than cultivated soybean as assessed by plant biomass paraments, total carbon content and total nitrogen content. Gas-exchange coefficients and chlorophylls contents maintained relatively stable in wild soybean young leaves, but opposite in cultivated soybean. Wild soybean young leaves also increased the transport of beneficial ions, such as B3+, Fe3+, Mn2+, H2PO4- and C2O42-. In wild soybean old leaves, the nitrogen metabolism pathway was significant enhanced, especially the aspartic acid and GABA metabolisms. While in cultivated soybean, the nitrogen metabolism decreased obviously in young leaves but had no significant change in old leaves. The phenylpropanoid metabolism pathway was also activated in wild soybean. Contrary to cultivated soybeans, wild soybean tricarboxylic acid cycle and carbon metabolism including polyols and organic acids consolidated in old leaves and maintained a relative normal state in young leaves. These strategies could improve the antioxidant and N-fixation capacity in wild soybean. CONCLUSION: The survival and growth of wild soybean under low nitrogen stress conditions relied on physiological adjustments and metabolic changes that occurred at the cellular level. Compared with cultivated soybean, wild soybean young leaves could maintain a relatively normal growth mainly owing to a significant enhancement of key amino acids and nonprotein nitrogen metabolism in old leaves, especially aspartic acid, proline metabolism which provided basis for nitrogen reutilization from old leaves to young leaves. Consolidating the tricarboxylic acid cycle, intensifying phenylpropanoid metabolism, and accumulating more polyols and organic acids also had positive effect on it.

Ionomic and metabolic responses to neutral salt or alkaline salt stresses in maize (Zea mays L.) seedlings.

BACKGROUND: Soil salinity and alkalinity present a serious threat to global agriculture. However, most of the studies have focused on neutral salt stress, and the information on the metabolic responses of plants to alkaline salt stress is limited. This investigation aimed at determining the influence of neutral salt and alkaline salt stresses on the content of metal elements and metabolites in maize plant tissues, by using mixtures of various proportions of NaCl, NaHCO3, Na2SO4, and Na2CO3. RESULTS: We found that alkaline salt stress suppressed more pronouncedly the photosynthesis and growth of maize plants than salinity stress. Under alkaline salt stress conditions, metal ions formed massive precipitates, which ultimately reduced plant nutrient availability. On the other hand, high neutral salt stress induced metabolic changes in the direction of gluconeogenesis leading to the enhanced formation of sugars as a reaction contributing to the mitigation of osmotic stress. Thus, the active synthesis of sugars in shoots was essential to the development of salt tolerance. However, the alkaline salt stress conditions characterized by elevated pH values suppressed substantially the levels of photosynthesis, N metabolism, glycolysis, and the production of sugars and amino acids. CONCLUSIONS: These results indicate the presence of different defensive mechanisms responsible for the plant responses to neutral salt and alkaline salt stresses. In addition, the increased concentration of organic acids and enhanced metabolic energy might be potential major factors that can contribute to the maintenance intracellular ion balance in maize plants and counteract the negative effects of high pH under alkaline salt stress.

Cloning and functional analysis of the promoters that upregulate carotenogenic gene expression during flower development in Gentiana lutea.

Over the last two decades, many carotenogenic genes have been cloned and used to generate metabolically engineered plants producing higher levels of carotenoids. However, comparatively little is known about the regulation of endogenous carotenogenic genes in higher plants, and this restricts our ability to predict how engineered plants will perform in terms of carotenoid content and composition. During petal development in the Great Yellow Gentian (Gentiana lutea), carotenoid accumulation, the formation of chromoplasts and the upregulation of several carotenogenic genes are temporally coordinated. We investigated the regulatory mechanisms responsible for this coordinated expression by isolating five G. lutea carotenogenic gene (GlPDS, GlZDS, GlLYCB, GlBCH and GlLYCE) promoters by inverse polymerase chain reaction (PCR). Each promoter was sufficient for developmentally regulated expression of the gusA reporter gene following transient expression in tomato (Solanum lycopersicum cv. Micro-Tom). Interestingly, the GlLYCB and GlBCH promoters drove high levels of gusA expression in chromoplast-containing mature green fruits, but low levels in chloroplast-containing immature green fruits, indicating a strict correlation between promoter activity, tomato fruit development and chromoplast differentiation. As well as core promoter elements such as TATA and CAAT boxes, all five promoters together with previously characterized GlZEP promoter contained three common cis-regulatory motifs involved in the response to methyl jasmonate (CGTCA) and ethylene (ATCTA), and required for endosperm expression (Skn-1_motif, GTCAT). These shared common cis-acting elements may represent binding sites for transcription factors responsible for co-regulation. Our data provide insight into the regulatory basis of the coordinated upregulation of carotenogenic gene expression during flower development in G. lutea.

Comparative differences in maintaining membrane fluidity and remodeling cell wall between Glycine soja and Glycine max leaves under drought.

Water shortage is one of the most important environmental factors limiting crop yield. In this study, we used wild soybean (Glycine soja Sieb. et Zucc.) and soybean (Glycinemax (L.) Merr.) seedlings as experimental materials, simulated drought stress using soil gravimetry, measured growth and physiological parameters, and analyzed differentially expressed genes and metabolites in the leaves of seedling by integrated transcriptomics and metabolomics techniques. The results indicate that under water deficit, Glycine soja maintained stable photosynthate by accumulating Mg2+, Fe3+, Mn2+, Zn2+ and B3+, and improved water absorption by increasing root growth. Notably, Glycine soja enhanced linoleic acid metabolism and plasma membrane intrinsic protein (PIP1) gene expression to maintain membrane fluidity, and increased pentose, glucuronate and galactose metabolism and thaumatin protein genes expression to remodel the cell wall, thereby increasing water-absorption to better tolerate to drought stress. In addition, it was found that secondary phenolic metabolism, such as phenylpropane biosynthesis, flavonoid biosynthesis and ascobate and aldarate metabolism were weakened, resulting in the collapse of the antioxidant system, which was the main reason for the sensitivity of Glycine max to drought stress. These results provide new insights into plant adaptation to water deficit and offer a theoretical basis for breeding soybean varieties with drought tolerance.

Comparative study of leaf nutrient reabsorption by two different ecotypes of wild soybean under low-nitrogen stress.

Wild soybean (Glycine soja), the ancestor of cultivated soybean, has evolved into many ecotypes with different adaptations to adversity under the action of divergent evolution. Barren-tolerant wild soybean has developed adaptation to most nutrient-stress environments, especially with respect to low nitrogen (LN) conditions. This study describes the differences in physiological and metabolomic changes between common wild soybean (GS1) and barren-tolerant wild soybean(GS2) under LN stress. Compared with plants grown under the unstressed control (CK) conditions, the young leaves of barren-tolerant wild soybean under LN conditions maintained relatively stable chlorophyll, concentration and rates of photosynthesis and transpiration, as well as increased carotenoid content, whereas the net photosynthetic rate (PN) of GS1 decreased significantly 0.64-fold (p < 0.05) in the young leaves of GS1. The ratio of internal to atmospheric CO2 concentrations increased significantly 0.07-fold (p < 0.05), 0.09-fold (p < 0.05) in the young leaves of GS1 and GS2, respectively, and increased significantly 0.05-fold (p < 0.05) and 0.07-fold (p < 0.05) in the old leaves of GS1 and GS2, respectively, relative to the CK. The concentration of chlorophylls a and b decreased significantly 0.45-fold (p < 0.05), 0.13-fold (p > 0.05) in the young leaves of GS1 and GS2, respectively, and decreased significantly 0.74-fold (p < 0.01) and 0.60-fold (p < 0.01) in the old leaves of GS1 and GS2, respectively. Under LN stress, nitrate concentration in the young leaves of GS1 and GS2 decreased significantly 0.69- and 0.50-fold (p < 0.01), respectively, relative to CK, and decreased significantly 2.10-fold and 1.77-fold (p < 0.01) in the old leaves of GS1 and GS2, respectively. Barren-tolerant wild soybean increased the concentration of beneficial ion pairs. Under LN stress, Zn2+ significantly increased by 1.06- and 1.35-fold (p < 0.01) in the young and old leaves of GS2 (p < 0.01), but there was no significant change in GS1. The metabolism of amino acids and organic acids was high in GS2 young and old leaves, and the metabolites related to the TCA cycle were significantly increased. The 4-aminobutyric acid (GABA) concertation decreased significantly 0.70-fold (p < 0.05) in the young leaves of GS1 but increased 0.21-fold (p < 0.05) significantly in GS2. The relative concentration of proline increased significantly 1.21-fold (p < 0.01) and 2.85-fold (p < 0.01) in the young and old leaves of GS2. Under LN stress, GS2 could maintain photosynthesis rate and enhance the reabsorption of nitrate and magnesium in young leaves, compared to GS1. More importantly, GS2 exhibited increased amino acid and TCA cycle metabolism in young and old leaves. Adequate reabsorption of mineral and organic nutrients is an important strategy for barren-tolerant wild soybeans to survive under LN stress. Our research provides a new perspective on the exploitation and utilization of wild soybean resources.

Integration of the transcriptome and metabolome reveals the mechanism of resistance to low phosphorus in wild soybean seedling leaves.

Plant growth, development, yield and quality are limited by barren soil. Soil phosphorus deficiency is one of the common factors causing soil barrenness. Plants have evolved morphological, physiological and molecular adaptations to resist to phosphorus deficiency. Wild soybean, a wild relative of cultivated soybean, has an obvious genetic relationship with cultivated soybean and has many beneficial characteristics such as strong low phosphorus resistance. Therefore, in this study, the integration analysis of transcriptome and metabolome of wild and cultivated soybean seedlings leaves were applied under phosphorus deficiency to reveal the mechanism of resistance to low phosphorus stress in wild soybean leaves, especially the key role of membrane phospholipid reuse and protection. Under phosphorus deficiency, wild soybean resisted low phosphorus stress by enhancing phosphorus reuse and strengthening membrane protection mechanisms, that is, by enhancing phospholipid metabolism, degrading membrane phospholipids, releasing phosphorus, increasing phosphorus reuse, and enhancing galactolipid biosynthesis. This, in turn, produced digalactosyl diacylglycerol to replace missing phospholipids for membrane maintenance and enhanced glutathione metabolism to protect the membrane system from damage. At the same time, phosphorus deficiency increased the levels of the intermediate metabolites glycine and ornithine, while significantly regulating the expression of transcription factors WRKY75 and MYB86. The enhancement of these metabolic pathways and the significant regulation of gene expression play an important role in improving the low phosphorus tolerance of wild soybean. This study will provide a useful theoretical basis for breeding soybean with low phosphorus tolerance.

Functional characterization of the Gentiana lutea zeaxanthin epoxidase (GlZEP) promoter in transgenic tomato plants.

The accumulation of carotenoids in plants depends critically on the spatiotemporal expression profiles of the genes encoding enzymes in the carotenogenic pathway. We cloned and characterized the Gentiana lutea zeaxanthin epoxidase (GlZEP) promoter to determine its role in the regulation of carotenogenesis, because the native gene is expressed at high levels in petals, which contain abundant chromoplasts. We transformed tomato (Solanum lycopersicum cv. Micro-Tom) plants with the gusA gene encoding the reporter enzyme ß-glucuronidase (GUS) under the control of the GlZEP promoter, and investigated the reporter expression profile at the mRNA and protein levels. We detected high levels of gusA expression and GUS activity in chromoplast-containing flowers and fruits, but minimal levels in immature fruits containing green chloroplasts, in sepals, leaves, stems and roots. GlZEP-gusA expression was strictly associated with fruit development and chromoplast differentiation, suggesting an evolutionarily-conserved link between ZEP and the differentiation of organelles that store carotenoid pigments. The impact of our results on current models for the regulation of carotenogenesis in plants is discussed.

Integrating transcriptomic and metabolomic analysis in roots of wild soybean seedlings in response to low-phosphorus stress.

Introduction: Plants undergo divergent adaptations to form different ecotypes when exposed to different habitats. Ecotypes with ecological adaptation advantages are excellent germplasm resources for crop improvement. Methods: his study comprehensively compared the differences in morphology and physiological mechanisms in the roots of two different ecotypes of wild soybean (Glycine soja) seedlings under artificially simulated low-phosphorus (LP) stress. Result: The seedlings of barren-tolerant wild soybean (GS2) suffered less damage than common wild soybean (GS1). GS2 absorbed more phosphorus (P) by increasing root length. In-depth integrated analyses of transcriptomics and metabolomics revealed the formation process of the ecological adaptability of the two different ecotypes wild soybean from the perspective of gene expression and metabolic changes. This study revealed the adaptation process of GS2 from the perspective of the adaptation of structural and molecular metabolism, mainly including: (1) Enhancing the metabolism of phenolic compounds, lignin, and organic acid metabolism could activate unavailable soil P; (2) Up-regulating genes encoding pectinesterase and phospholipase C (PLC) specifically could promote the reuse of structural P; (3) Some factors could reduce the oxidative damage to the membranes caused by LP stress, such as accumulating the metabolites putrescine and ascorbate significantly, up-regulating the genes encoding SQD2 (the key enzyme of sulfolipid substitution of phospholipids) substantially and enhancing the synthesis of secondary antioxidant metabolite anthocyanins and the AsA-GSH cycle; (4) enhancing the uptake of soil P by upregulating inorganic phosphate transporter, acid phosphatase ACP1, and purple acid phosphatase genes; (5) HSFA6b and MYB61 are the key TFs to resist LP stress. Discussion: In general, GS2 could resist LP stress by activating unavailable soil P, reusing plant structural P, rebuilding membrane lipids, and enhancing the antioxidant membrane protection system. Our study provides a new perspective for the study of divergent adaptation of plants.

The Coordinated Upregulated Expression of Genes Involved in MEP, Chlorophyll, Carotenoid and Tocopherol Pathways, Mirrored the Corresponding Metabolite Contents in Rice Leaves during De-Etiolation.

Light is an essential regulator of many developmental processes in higher plants. We investigated the effect of 4-hydroxy-3-methylbut-2-enyl diphosphate reductase 1/2 genes (OsHDR1/2) and isopentenyl diphosphate isomerase 1/2 genes (OsIPPI1/2) on the biosynthesis of chlorophylls, carotenoids, and phytosterols in 14-day-old etiolated rice (Oyza sativa L.) leaves during de-etiolation. However, little is known about the effect of isoprenoid biosynthesis genes on the corresponding metabolites during the de-etiolation of etiolated rice leaves. The results showed that the levels of α-tocopherol were significantly increased in de-etiolated rice leaves. Similar to 1-deoxy-D-xylulose-5-phosphate synthase 3 gene (OsDXS3), both OsDXS1 and OsDXS2 genes encode functional 1-deoxy-D-xylulose-5-phosphate synthase (DXS) activities. Their expression patterns and the synthesis of chlorophyll, carotenoid, and tocopherol metabolites suggested that OsDXS1 is responsible for the biosynthesis of plastidial isoprenoids in de-etiolated rice leaves. The expression analysis of isoprenoid biosynthesis genes revealed that the coordinated expression of the MEP (2-C-methyl-D-erythritol 4-phosphate) pathway, chlorophyll, carotenoid, and tocopherol pathway genes mirrored the changes in the levels of the corresponding metabolites during de-etiolation. The underpinning mechanistic basis of coordinated light-upregulated gene expression was elucidated during the de-etiolation process, specifically the role of light-responsive cis-regulatory motifs in the promoter region of these genes. In silico promoter analysis showed that the light-responsive cis-regulatory elements presented in all the promoter regions of each light-upregulated gene, providing an important link between observed phenotype during de-etiolation and the molecular machinery controlling expression of these genes.

The regulation of carotenoid pigmentation in flowers.

Carotenoids fulfill many processes that are essential for normal growth and development in plants, but they are also responsible for the breathtaking variety of red-to-yellow colors we see in flowers and fruits. Although such visual diversity helps to attract pollinators and encourages herbivores to distribute seeds, humans also benefit from the aesthetic properties of flowers and an entire floriculture industry has developed on the basis that new and attractive varieties can be produced. Over the last decade, much has been learned about the impact of carotenoid metabolism on flower color development and the molecular basis of flower color. A number of different regulatory mechanisms have been described ranging from the transcriptional regulation of genes involved in carotenoid synthesis to the control of carotenoid storage in sink organs. This means we can now explain many of the natural colorful varieties we see around us and also engineer plants to produce flowers with novel and exciting varieties that are not provided by nature.

Comparative Study of the Effects of Salinity on Growth, Gas Exchange, N Accumulation and Stable Isotope Signatures of Forage Oat (Avena sativa L.) Genotypes.

Identifying suitable salt stress-tolerant phenotypes based on their agronomic and physiological traits remains a herculean task in forage-type oat (Avena sativa L.) breeding. This study examined the responses of six forage-type oat cultivars under four levels of saline stress over the vegetative growth cycle. Crop growth, water status-related traits and nitrogen status-related traits were analyzed in different plant parts to evaluate effective approaches for identifying salt tolerance. Plant biomass, height, tiller number and culm thickness changed substantially during salinity, but they were not precise enough for use in estimating genotypic salinity tolerance during long-term stress. Genotypes bearing larger numbers of tillers showed greater sensitivity to salinity due to its effects on biomass loss. Tolerant genotypes exhibited higher relative shoot biomass together with higher water use efficiency. The concentrations of Na+, K+ and their ratio, combined with the δ13C in shoots and roots were effective indicators for estimating tolerant genotypes through better water maintenance. N concentrations of shoots were the most efficient for evaluating genotypic tolerance. Low nitrate reductase (NR) and glutamine synthetase (GS) activity might be key factors limiting N accumulation. Chlorophyll (Chl) content and net photosynthetic rate, as well as stomatal conductance and evaporation, were useful for identifying salinity tolerance physiological mechanisms, but the effectiveness was low for genotypic tolerance testing for forage type oats due to the interaction between genotypes and salinity levels. The selection of high salinity-tolerant genotypes should focus on genotypes with photosynthetic resilience to salt, followed by high N metabolism (higher NR and GS activities) to ensure accumulation of more N in the shoot dry matter.

Effects of salinity on photosynthetic traits, ion homeostasis and nitrogen metabolism in wild and cultivated soybean.

BACKGROUND: Carbon and nitrogen metabolism need to be highly regulated to achieve cell acclimation to changing environmental conditions. The understanding of physio-biochemical responses of crops to salinity stress could help to stabilize their performance and yield. In this study we have analyzed the roles of photosynthesis, ion physiology and nitrate assimilation toward saline/alkaline stress acclimation in wild and cultivated soybean seedlings. METHODS: Growth and photosynthetic parameters, ion concentrations and the activity of enzymes involved in nitrogen assimilation were determined in seedlings of one wild and one cultivated soybean accession subjected to saline or alkaline stresses. RESULTS: Both saline and alkaline stresses had a negative impact on the growth and metabolism of both wild and cultivated soybean.The growth, photosynthesis, and gas exchange parameters showed a significant decrease in response to increasing salt concentration. Additionally, a significant increase in root Na+ and Cl- concentration was observed. However, photosynthetic performance and ion regulation were higher in wild than in cultivated soybean under saline and alkaline stresses. Nitrate reductase (NR) and the glutamine synthetase/glutamate synthase (GS/GOGAT) cycle showed a significant decrease in leaves of both genotypes. The reduction in the GS/GOGAT cycle was accompanied by high aminating glutamate dehydrogenase (NADH-glutamate dehydrogenase) activity, indicating the assimilation of high levels of NH4 +. A significant increase in the activities of aminating and deaminating enzymes, including glutamate dehydrogenase (GDH), alanine aminotransferase (AlaAT) and aspartate aminotransferase (AspAT), was observed, probably due to the high glutamate demand and maintenance of the Krebs cycle to correct the C: N status. CONCLUSIONS: Cultivated soybean was much more stress sensitive than was the wild soybean. The decrease in growth, photosynthesis, ion regulation and nitrogen assimilation enzymes was greater in cultivated soybean than in wild soybean. The impact of alkaline stress was more pronounced than that of saline stress. Wild soybean regulated the physiological mechanisms of photosynthesis and nitrate assimilation more effectively than did cultivated soybean. The present findings provide a theoretical basis with which to screen and utilize wild and cultivated soybean germplasm for breeding new stress-tolerant soybean.