Physiological and biochemical response analysis of Styrax tonkinensis seedlings to waterlogging stress.

Climate change is increasing flooding in provinces of the south of the Yangtze River, posing challenges for promoting Styrax tonkinensis seedlings in these areas. To understand the physiological reasons for this species' intolerance to waterlogging, we observed biochemical parameters in one-year-old S. tonkinensis seedlings during two seasons. For 4 and 12 days in summer and winter experiments, respectively, we subjected seedlings to a pot-in-pot waterlogging treatment. Control groups were established at 0 h and 0 days. We examined indicators related to root vigor, reactive oxygen species (ROS), antioxidant enzymes, fermentative pathways, and more. The results displayed that decreased abscisic acid accumulation in roots inhibited water transport. Increased dehydrogenase and lactate dehydrogenase activity in roots promoted alcohol and lactate fermentation, causing toxic damage and reduced root vigor, impeding water absorption. In leaves, high ROS levels led to lipid peroxidation, exacerbating water loss from continuous transpiration. The high relative electric conductivity and low leaf relative water content indicated water loss, causing leaf wilting and shriveling. Conversely, winter seedlings, devoid of leaves, significantly reduced transpiration, and dormancy delayed root fermentation. With less ROS damage in roots, winter seedlings exhibited greater waterlogging tolerance. In summary, excessive water loss from leaves and inhibited vertical water transport contributed to low summer survival rates, while winter leafless dormancy and reduced ROS damage enhanced tolerance. Our findings provide insights for enhancing waterlogging resistance in S. tonkinensis amidst climate change challenges.

The Combined Analysis of Transcriptome and Antioxidant Enzymes Revealed the Mechanism of EBL and ZnO NPs Enhancing Styrax tonkinensis Seed Abiotic Stress Resistance.

As global climate change worsens, trees will have difficulties adapting to abiotic pressures, particularly in the field, where environmental characteristics are difficult to control. A prospective commercial and ornamental tree species, Styrax tonkinensis, has its seed oil output and quality reduced as a result, which lowers the economic benefits. This necessitates growers to implement efficient strategies to increase the seeds of woody biofuel species' tolerance to abiotic stress. Numerous studies have shown that ZnO nanoparticles (NPs), a new material, and BRs assist plants to increase their resilience to abiotic stress and subsequently adapt to it. However, there have not been many investigations into S. tonkinensis seed resistance. In this study, we examined the changes in antioxidant enzyme activities and transcriptomic results of S. tonkinensis seeds throughout the seed development period to investigate the effects of 24-epibrassinolide (EBL), one of the BRs, and ZnO NPs treatments alone or together on the stress resistance of S. tonkinensis seeds. On 70, 100, and 130 days after flowering (DAF), spraying EBL or ZnO NPs increased the activity of antioxidant enzymes (POD, SOD, and CAT) in S. tonkinensis seeds. Moreover, when the EBL and ZnO NPs were sprayed together, the activities of antioxidant enzymes were the strongest, which suggests that the positive effects of the two can be superimposed. On 70 and 100 DAF, the EBL and ZnO NPs treatments improved seed stress resistance, mostly through complex plant hormone crosstalk signaling, which includes IAA, JA, BR, and ABA signaling. Additionally, ABA played an essential role in hormone crosstalk, while, on 130 DAF, due to the physiological characteristics of seeds themselves in the late stage of maturity, the improvement in seed stress resistance by EBL and ZnO NPs was related to protein synthesis, especially late embryogenesis-abundant protein (LEA), and other nutrient storage in seeds. Spraying EBL and ZnO NPs during the seed growth of S. tonkinensis could significantly increase seed stress resistance. Our findings provide fresh perspectives on how cultural practices can increase abiotic stress tolerance in woody seedlings.

Transcriptomic Analyses Reveal Key Genes Involved in Pigment Biosynthesis Related to Leaf Color Change of Liquidambar formosana Hance.

Liquidambar formosana Hance has a highly ornamental value as an important urban greening tree species with bright and beautiful leaf color. To gain insights into the physiological and molecular mechanisms of L. formosana leaf color change, the leaves of three different clones were sampled every ten days from October 13, 2019, five times in total, which are S1, S2, S3, S4 and S5. Transcriptome sequencing was performed at S1 and S4. The chlorophyll content of the three clones decreased significantly, while the anthocyanins content of the three clones increased significantly in the coloring stage. The anthocyanins content of clone 2 was far more than that of the other two clones throughout the period of leaf color change. The transcriptome analysis showed that six DEGs related to anthocyanins biosynthesis, including CHS (chalcone synthase), CHI (chalcone isomerase), F3'H (flavonoid 3'-hydroxylase), DFR (dihydroflavonol 4-reductase), ANS (anthocyanidin synthase) and FLS (flavonol synthase), were found in three clones. Clone 2 has another three DEGs related to anthocyanins biosynthesis, including PAL (Phenylalanine ammonia-lyase), F3'5'H (flavonoid 3',5'-hydroxylase) and UFGT (flavonoid 3-O-glucosyltransferase). We lay a foundation for understanding the molecular regulation mechanism of the formation of leaf color by exploring valuable genes, which is helpful for L. formosana breeding.

24-Epibrassinolide Promotes Fatty Acid Accumulation and the Expression of Related Genes in Styrax tonkinensis Seeds.

Styrax tonkinensis, whose seeds are rich in unsaturated fatty acids (UFAs), is a high oil value tree species, and the seed oil has perfect biodiesel properties. Therefore, the elucidation of the effect of 24-epibrassinolide (EBL) on fatty acid (FA) concentration and the expression of FA biosynthesis-related genes is critical for deeply studying the seed oil in S. tonkinensis. In this study, we aimed to investigate the changing trend of FA concentration and composition and identify candidate genes involved in FA biosynthesis under EBL treatment using transcriptome sequencing and GC-MS. The results showed that 5 µmol/L of EBL (EBL5) boosted the accumulation of FA and had the hugest effect on FA concentration at 70 days after flowering (DAF). A total of 20 FAs were identified; among them, palmitic acid, oleic acid, linoleic acid, and linolenic acid were the main components. In total, 117,904 unigenes were detected, and the average length was 1120 bp. Among them, 1205 unigenes were assigned to 'lipid translations and metabolism' in COG categories, while 290 unigenes were assigned to 'biosynthesis of unsaturated fatty acid' in KEGG categories. Twelve important genes related to FA biosynthesis were identified, and their expression levels were confirmed by quantitative real-time PCR. KAR, KASIII, and accA, encoding FA biosynthesis-related enzymes, all expressed the highest at 70 DAF, which was coincident with a rapid rise in FA concentration during seed development. FAD2 and FATB conduced to UFA and saturated fatty acids (SFA) accumulation, respectively. EBL5 induced the expression of FA biosynthesis-related genes. The concentration of FA was increased after EBL5 application, and EBL5 also enhanced the enzyme activity by promoting the expression of genes related to FA biosynthesis. Our research could provide a reference for understanding the FA biosynthesis of S. tonkinensis seeds at physiological and molecular levels.

Transcriptomic and Metabolomic Analysis Unravels the Molecular Regulatory Mechanism of Fatty Acid Biosynthesis in Styrax tonkinensis Seeds under Methyl Jasmonate Treatment.

As the germ of a highly productive oil tree species, Styrax tonkinensis seeds have great potential to produce biodiesel and they have marvelous fatty acid (FA) composition. In order to explore the molecular regulatory mechanism of FA biosynthesis in S. tonkinensis seeds after methyl jasmonate (MJ) application, transcriptomic and metabolomic techniques were adopted so as to dissect the genes that are related to FA biosynthesis and their expression levels, as well as to discover the major FA concentration and composition. The results revealed that 200 µmol/L of MJ (MJ200) increased the crude fat (CF) mass fraction and generated the greatest impact on CF accumulation at 70 days after flowering. Twenty FAs were identified, among which palmitic acid, oleic acid, linoleic acid and linolenic acid were the major FAs, and the presence of MJ200 affected their concentrations variously. MJ200 could enhance FA accumulation through elevating the activity of enzymes that are related to FA synthesis. The number of differentially expressed genes increased with the seeds' development in general. Fatty acid biosynthesis, the biosynthesis of unsaturated fatty acid, fatty acid elongation and glycerolipid metabolism were the main lipid metabolism pathways that were found to be involved. The changes in the expression levels of EAR, KAR, accA, accB and SAD2 were consistent with the changes in the CF mass fraction, indicating that they are important genes in the FA biosynthesis of S. tonkinensis seeds and that MJ200 promoted their expression levels. In addition, bZIP (which was screened by weighted correlation network analysis) also created significant impacts on FA biosynthesis. Our research has provided a basis for further studies on FA biosynthesis that is regulated by MJ200 at the molecular level and has helped to clarify the functions of key genes in the FA metabolic pathway in S. tonkinensis seeds.