Identification and characterization of the karrikins signaling gene SsSMAX1 in Sapium sebiferum.

SUPPRESSOR OF MAX2 LIKE 1 (SMAX1) is a member of the SUPPRESSOR of MAX2 1­LIKE family of genes and is known as a target protein of KARRIKIN INSENSITIVE2 (KAI2)-MORE AXILLARY BRANCHES2 (MAX2), which mediates karrikin signaling in Arabidopsis. SMAX1 plays a significant role in seed germination, hypocotyl elongation, and root hair development in Arabidopsis. SMAX1 has not yet been identified and characterized in woody plants. This study identified and characterized SsSMAX1 in Sapium sebiferum and found that SsSMAX1 was highly expressed in the seed, hypocotyl, and root tips of S. sebiferum. SsSMAX1 was functionally characterized by ectopic expression in Arabidopsis. SsSMAX1 overexpression lines of Arabidopsis showed significantly delayed seed germination and produced seedlings with longer hypocotyl and roots than wild-type and Atsmax1 functional mutants. SsSMAX1 overexpression lines of Arabidopsis also had broader and longer leaves and petioles than wild-type and Atsmax1, suggesting that SsSMAX1 is functionally conserved. This study characterizes the SMAX1 gene in a woody and commercially valuable bioenergy plant, Sapium sebiferum. The results of this study are beneficial to future research on the molecular biology of woody plants.

Karrikinolide alleviates salt stress in wheat by regulating the redox and K+/Na+ homeostasis.

Karrikinolide (KAR1), identified in biochars, has gained research attention because of its significant role in seed germination, seedling development, root development, and abiotic stresses. However, KAR1 regulation of salt stress in wheat is elusive. This study investigated the physiological mechanism involved in KAR1 alleviation of salt stress in wheat. The results showed KAR1 boosted seed germination percentage under salinity stress via stimulating the relative expression of genes regulating gibberellins biosynthesis and decreasing the expression levels of abscisic acid biosynthesis and signaling genes. As seen in seed germination, exogenous supplementation of KAR1 dramatically mitigated the salt stress also in wheat seedling, resulting in increased root and shoot growth as measured in biomass as compared to salt stress alone. Salt stress significantly induced the endogenous hydrogen peroxide and malondialdehyde levels, whereas KAR1 strictly counterbalanced them. Under salt stress, KAR1 supplementation showed significant induction in reduced glutathione (GSH) and reduction in oxidized glutathione (GSSG) content, which improved GSH/GSSG ratio in wheat seedlings. Exogenous supplementation of KAR1 significantly promoted the activities of enzymatic antioxidants in wheat seedlings exposed to salt stress. KAR1 induced the relative expression of genes regulating the biosynthesis of antioxidants in wheat seedlings under salinity. Moreover, KAR1 induced the expression level of K+/Na+ homeostasis genes, reduced Na+ concentration, and induced K+ concentration in wheat seedling under salt stress. The results suggest that KAR1 supplementation maintained the redox and K+/Na+ homeostasis in wheat seedling under salinity, which might be a crucial part of physiological mechanisms in KAR1 induced tolerance to salt stress. In conclusion, we exposed the protective role of KAR1 against salt stress in wheat.

Overexpression of Karrikins Receptor Gene Sapium sebiferum KAI2 Promotes the Cold Stress Tolerance via Regulating the Redox Homeostasis in Arabidopsis thaliana.

KARRIKINS INSENSITIVE2 (KAI2) is the receptor gene for karrikins, recently found to be involved in seed germination, hypocotyl development, and the alleviation of salinity and osmotic stresses. Nevertheless, whether KAI2 could regulate cold tolerance remains elusive. In the present study, we identified that Arabidopsis mutants of KAI2 had a high mortality rate, while overexpression of, a bioenergy plant, Sapium sebiferum KAI2 (SsKAI2) significantly recovered the plants after cold stress. The results showed that the SsKAI2 overexpression lines (OEs) had significantly increased levels of proline, total soluble sugars, and total soluble protein. Meanwhile, SsKAI2 OEs had a much higher expression of cold-stress-acclimation-relate genes, such as Cold Shock Proteins and C-REPEAT BINDING FACTORS under cold stress. Moreover, the results showed that SsKAI2 OEs were hypersensitive to abscisic acid (ABA), and ABA signaling genes were w significantly affected in SsKAI2 OEs under cold stress, suggesting a potential interaction between SsKAI2 and ABA downstream signaling. In SsKAI2 OEs, the electrolyte leakage, hydrogen peroxide, and malondialdehyde contents were reduced under cold stress in Arabidopsis. SsKAI2 OEs enhanced the anti-oxidants like ascorbate peroxidase, catalase, peroxidase, superoxide dismutase, and total glutathione level under cold stress. Conclusively, these results provide novel insights into the understanding of karrikins role in the regulation of cold stress adaptation.

Karrikin Improves Osmotic and Salt Stress Tolerance via the Regulation of the Redox Homeostasis in the Oil Plant Sapium sebiferum.

Karrikins are reported to stimulate seed germination, regulate seedling growth, and increase the seedling vigor in abiotic stress conditions in plants. Nevertheless, how karrikins alleviate abiotic stress remains largely elusive. In this study, we found that karrikin (KAR1) could significantly alleviate both drought and salt stress in the important oil plant Sapium sebiferum. KAR1 supplementation in growth medium at a nanomolar (nM) concentration was enough to recover seed germination under salt and osmotic stress conditions. One nanomolar of KAR1 improved seedling biomass, increased the taproot length, and increased the number of lateral roots under abiotic stresses, suggesting that KAR1 is a potent alleviator of abiotic stresses in plants. Under abiotic stresses, KAR1-treated seedlings had a higher activity of the key antioxidative enzymes, such as superoxide dismutase, peroxidase, catalase, and ascorbate peroxidase, in comparison with the control, which leads to a lower level of hydrogen peroxide, malondialdehyde, and electrolyte leakage. Moreover, the metabolome analysis showed that KAR1 treatment significantly increased the level of organic acids and amino acids, which played important roles in redox homeostasis under stresses, suggesting that karrikins might alleviate abiotic stresses via the regulation of redox homeostasis. Under abiotic stresses, applications of karrikins did not increase the endogenous abscisic acid level but altered the expression of several ABA signaling genes, such as SNF1-RELATED PROTEIN KINASE2.3, SNF1-RELATED PROTEIN KINASE2.6, ABI3, and ABI5, suggesting potential interactions between karrikins and ABA signaling in the stress responses. Conclusively, we not only provided the physiological and molecular evidence to clarify the mechanism of karrikins in the regulation of stress adaptation in S. sebiferum but also showed the potential value of karrikins in agricultural practices, which will lay a foundation for further studies about the role of karrikins in abiotic stress alleviation in plants.

Genome-wide identification, characterization and expression pattern analysis of APYRASE family members in response to abiotic and biotic stresses in wheat.

APYRASEs, which directly regulate intra- and extra-cellular ATP homeostasis, play a pivotal role in the regulation of various stress adaptations in mammals, bacteria and plants. In the present study, we identified and characterized wheat APYRASE family members at the genomic level in wheat. The results identified a total of nine APY homologs with conserved ACR domains. The sequence alignments, phylogenetic relations and conserved motifs of wheat APYs were bioinformatically analyzed. Although they share highly conserved secondary and tertiary structures, the wheat APYs could be mainly categorized into three groups, according to phylogenetic and structural analysis. Additionally, these APYs exhibited similar expression patterns in the root and shoot, among which TaAPY3-1, TaAPY3-3 and TaAPY3-4 had the highest expression levels. The time-course expression patterns of the eight APYs in response to biotic and abiotic stress in the wheat seedlings were also investigated. TaAPY3-2, TaAPY3-3, TaAPY3-4 and TaAPY6 exhibited strong sensitivity to all kinds of stresses in the leaves. Some APYs showed specific expression responses, such as TaAPY6 to heavy metal stress, and TaAPY7 to heat and salt stress. These results suggest that the stress-inducible APYs could have potential roles in the regulation of environmental stress adaptations. Moreover, the catalytic activity of TaAPY3-1 was further analyzed in the in vitro system. The results showed that TaAPY3-1 protein exhibited high catalytic activity in the degradation of ATP and ADP, but with low activity in degradation of TTP and GTP. It also has an extensive range of temperature adaptability, but preferred relatively acidic pH conditions. In this study, the genome-wide identification and characterization of APYs in wheat were suggested to be useful for further genetic modifications in the generation of high-stress-tolerant wheat cultivars.

iTRAQ-based quantitative proteome analysis reveals metabolic changes between a cleistogamous wheat mutant and its wild-type wheat counterpart.

BACKGROUND: Wheat is one of the most important staple crops worldwide. Fusarium head blight (FHB) severely affects wheat yield and quality. A novel bread wheat mutant, ZK001, characterized as cleistogamic was isolated from a non-cleistogamous variety Yumai 18 (YM18) through static magnetic field mutagenesis. Cleistogamy is a promising strategy for controlling FHB. However, little is known about the mechanism of cleistogamy in wheat. METHODS: We performed a FHB resistance test to identify the FHB infection rate of ZK001. We also measured the agronomic traits of ZK001 and the starch and total soluble sugar contents of lodicules in YM18 and ZK001. Finally, we performed comparative studies at the proteome level between YM18 and ZK001 based on the proteomic technique of isobaric tags for relative and absolute quantification. RESULTS: The infection rate of ZK001 was lower than that of its wild-type and Aikang 58. The abnormal lodicules of ZK001 lost the ability to push the lemma and palea apart during the flowering stage. Proteome analysis showed that the main differentially abundant proteins (DAPs) were related to carbohydrate metabolism, protein transport, and calcium ion binding. These DAPs may work together to regulate cellular homeostasis, osmotic pressure and the development of lodicules. This hypothesis is supported by the analysis of starch, soluble sugar content in the lodicules as well as the results of Quantitative reverse transcription polymerase chain reaction. CONCLUSIONS: Proteomic analysis has provided comprehensive information that should be useful for further research on the lodicule development mechanism in wheat. The ZK001 mutant is optimal for studying flower development in wheat and could be very important for FHB resistant projects via conventional crossing.

Comparative transcriptome analysis reveals the regulatory networks of cytokinin in promoting the floral feminization in the oil plant Sapium sebiferum.

BACKGROUND: Sapium sebiferum, whose seeds contain high level of fatty acids, has been considered as one of the most important oil plants. However, the high male to female flower ratio limited the seed yield improvement and its industrial potentials. Thus, the study of the sex determination in S. sebiferum is of significant importance in increasing the seed yield. RESULTS: In this study, we demonstrated that in S. sebiferum, cytokinin (CK) had strong feminization effects on the floral development. Exogenous application with 6-benzylaminopurine (6-BA) or thidiazuron (TDZ) significantly induced the development of female flowers and increased the fruit number. Interestingly, the feminization effects of cytokinin were also detected on the androecious genotype of S. sebiferum which only produce male flowers. To further investigate the mechanism underlying the role of cytokinin in the flower development and sex differentiation, we performed the comparative transcriptome analysis of the floral buds of the androecious plants subjected to 6-BA. The results showed that there were separately 129, 352 and 642 genes differentially expressed at 6 h, 12 h and 24 h after 6-BA treatment. Functional analysis of the differentially expressed genes (DEGs) showed that many genes are related to the hormonal biosynthesis and signaling, nutrients translocation and cell cycle. Moreover, there were twenty one flowering-related genes identified to be differentially regulated by 6-BA treatment. Specifically, the gynoecium development-related genes SPATULA (SPT), KANADI 2 (KAN2), JAGGED (JAG) and Cytochrome P450 78A9 (CYP79A9) were significantly up-regulated, whereas the expression of PISTILLATA (PI), TATA Box Associated Factor II 59 (TAFII59) and MYB Domain Protein 108 (MYB108) that were important for male organ development was down-regulated in response to 6-BA treatment, demonstrating that cytokinin could directly target the floral organ identity genes to regulate the flower sex. CONCLUSIONS: Our work demonstrated that cytokinin is a potential regulator in female flower development in S. sebiferum. The transcriptome analysis of the floral sex transition from androecious to monoecious in response to cytokinin treatment on the androecious S. sebiferum provided valuable information related to the mechanism of sex determination in the perennial woody plants.

Proanthocyanidins in seed coat tegmen and endospermic cap inhibit seed germination in Sapium sebiferum.

Sapium sebiferum, an ornamental and bio-energetic plant, is propagated by seed. Its seed coat contains germination inhibitors and takes a long time to stratify for germination. In this study, we discovered that the S. sebiferum seed coat (especially the tegmen) and endospermic cap (ESC) contained high levels of proanthocyanidins (PAs). Seed coat and ESC removal induced seed germination, whereas exogenous application with seed coat extract (SCE) or PAs significantly inhibited this process, suggesting that PAs in the seed coat played a major role in regulating seed germination in S. sebiferum. We further investigated how SCE affected the expression of the seed-germination-related genes. The results showed that treatment with SCE upregulated the transcription level of the dormancy-related gene, gibberellins (GAs) suppressing genes, abscisic acid (ABA) biosynthesis and signalling genes. SCE decreased the transcript levels of ABA catabolic genes, GAs biosynthesis genes, reactive oxygen species genes and nitrates-signalling genes. Exogenous application of nordihydroguaiaretic acid, gibberellic acid, hydrogen peroxide and potassium nitrate recovered seed germination in seed-coat-extract supplemented medium. In this study, we highlighted the role of PAs, and their interactions with the other germination regulators, in the regulation of seed dormancy in S. sebiferum.

Transcriptome analysis of pecan seeds at different developing stages and identification of key genes involved in lipid metabolism.

Pecan is an economically important nut crop tree due to its unique texture and flavor properties. The pecan seed is rich of unsaturated fatty acid and protein. However, little is known about the molecular mechanisms of the biosynthesis of fatty acids in the developing seeds. In this study, transcriptome sequencing of the developing seeds was performed using Illumina sequencing technology. Pecan seed embryos at different developmental stages were collected and sequenced. The transcriptomes of pecan seeds at two key developing stages (PA, the initial stage and PS, the fast oil accumulation stage) were also compared. A total of 82,155 unigenes, with an average length of 1,198 bp from seven independent libraries were generated. After functional annotations, we detected approximately 55,854 CDS, among which, 2,807 were Transcription Factor (TF) coding unigenes. Further, there were 13,325 unigenes that showed a 2-fold or greater expression difference between the two groups of libraries (two developmental stages). After transcriptome analysis, we identified abundant unigenes that could be involved in fatty acid biosynthesis, degradation and some other aspects of seed development in pecan. This study presents a comprehensive dataset of transcriptomic changes during the seed development of pecan. It provides insights in understanding the molecular mechanisms responsible for fatty acid biosynthesis in the seed development. The identification of functional genes will also be useful for the molecular breeding work of pecan.

Exogenous Melatonin Confers Cadmium Tolerance by Counterbalancing the Hydrogen Peroxide Homeostasis in Wheat Seedlings.

Melatonin has emerged as a research highlight regarding its important role in regulating plant growth and the adaptation to the environmental stresses. In this study, we investigated how melatonin prevented the cadmium toxicity to wheat seedlings. The results demonstrated that cadmium induced the expression of melatonin biosynthesis-related genes and cause a significant increase of endogenous melatonin level. Melatonin treatment drastically alleviated the cadmium toxicity, resulting in increased plant height, biomass accumulation, and root growth. Cadmium and senescence treatment significantly increased the endogenous level of hydrogen peroxide, which was strictly counterbalanced by melatonin. Furthermore, melatonin treatment caused a significant increase of GSH (reduced glutathione) content and the GSH/GSSG (oxidized glutathione) ratio. The activities of two key antioxidant enzymes, ascorbate peroxidase (APX) and superoxide dismutase (SOD), but not catalase (CAT) and peroxidase (POD), were specifically improved by melatonin. Additionally, melatonin not only promoted the primary root growth, but also drastically enhanced the capacity of the seedling roots to degrade the exogenous hydrogen peroxide. These results suggested that melatonin played a key role in maintaining the hydrogen peroxide homeostasis, via regulation of the antioxidant systems. Conclusively, this study revealed a crucial protective role of melatonin in the regulation of cadmium resistance in wheat.