Crosstalk between brassinosteroids and other phytohormones during plant development and stress adaption.

Brassinosteroids (BRs) are a group of polyhydroxylated phytosterols that play essential roles in regulating plant growth and development as well as stress adaptation. It is worth noting that BRs do not function alone, but rather they crosstalk with other endogenous signaling molecules, including the phytohormones auxin, cytokinins (CKs), gibberellins (GAs), abscisic acid (ABA), ethylene (ET), jasmonates (JAs), salicylic acid (SA), and strigolactones (SLs), forming elaborate signaling networks to modulate plant growth and development. BRs interact with other phytohormones mainly by regulating each others' homeostasis, transport, or signaling pathway at the transcriptional and posttranslational levels. In this review, we focus our attention on current research progress in BR signal transduction and the crosstalk between BRs and other phytohormones.

Exogenous brassinolide treatment regulates phenolic accumulation in mung bean sprouts through the modulation of sugar and energy metabolism.

BACKGROUND: The effects of exogenous brassinolide (BR) treatment (3.0 µmol L-1 ) on phenolic biosynthesis in mung bean sprouts were investigated. This investigation included the analysis of sugar content, substrates within the phenylpropane pathway, energy substances, enzymatic activity within the phenylpropane pathway, sugar metabolism and energy metabolism. RESULTS: Results showed that BR treatment significantly increased the levels of total phenolics, p-hydroxybenzoic acid, p-coumaric acid, gallic acid, fumalic acid and caffeic acid. This enhancement was accomplished through the elevation of l-phenylalanine levels and the activation of enzymes associated with the phenylpropane pathway in mung bean sprouts, including phenylalanine ammonia-lyase, cinnamate 4-hydroxylase and 4-coumarate CoA ligase. Furthermore, BR treatment induced alterations in sugar metabolism in mung bean sprouts as evidenced by the increased levels of glucose, fructose, sucrose and phosphoenolpyruvate. Moreover, increased activity was observed for enzymes linked to sucrose metabolism and glycolysis in the BR-treated group. Concurrently, BR treatment bolstered the levels of adenosine triphosphate and energy charge in mung bean sprouts, which was attributed to the activation of H+ -adenosine triphosphatase, Ca2+ -adenosine triphosphatase and succinic dehydrogenase. CONCLUSION: These results suggest that BR treatment can accelerate the accumulation of phenolic compounds in mung bean sprouts. This effect is achieved not only through the activation of the phenylpropane pathway, but also through the modulation of sugar and energy metabolism. The modulation provides ample energy and a substrate for the biosynthesis of phenolics. © 2023 Society of Chemical Industry.

A mutation in the promoter of the arabinogalactan protein 7-like gene PcAGP7-1 affects cell morphogenesis and brassinolide content in pear (Pyrus communis L.) stems.

Dwarfing rootstocks and dwarf cultivars are urgently needed for modern pear cultivation. However, germplasm resources for dwarfing pear are limited, and the underlying mechanisms remain unclear. We previously showed that dwarfism in pear is controlled by the single dominant gene PcDw (Dwarf). We report here that the expression of PcAGP7-1 (ARABINOGALACTAN PROTEIN 7-1), a key candidate gene for PcDw, is significantly higher in dwarf-type pear plants because of a mutation in an E-box in the promoter. Electrophoretic mobility shift assays and transient infiltration showed that the transcription factors PcBZR1 and PcBZR2 could directly bind to the E-box of the PcAGP7-1 promoter and repress transcription. Moreover, transgenic pear lines overexpressing PcAGP7-1 exhibited obvious dwarf phenotypes, whereas RNA interference pear lines for PcAGP7-1 were taller than controls. PcAGP7-1 overexpression also enhanced cell wall thickness, affected cell morphogenesis, and reduced brassinolide (BL) content, which inhibited BR signaling via a negative feedback loop, resulting in further dwarfing. Overall, we identified a dwarfing mechanism in perennial woody plants involving the BL-BZR/BES-AGP-BL regulatory module. Our findings provide insight into the molecular mechanism of plant dwarfism and suggest strategies for the molecular breeding of dwarf pear cultivars.

External application of brassinolide enhances cold resistance of tea plants (Camellia sinensis L.) by integrating calcium signals.

MAIN CONCLUSION: Exogenous brassinolide can activate the expression of key genes in the calcium signalling pathway to enhance cold resistance of tea plants. Brassinolide is an endogenous sterol phytohormone containing multiple hydroxyl groups that has the important function of improving plant cold resistance and alleviating freeze damage. To explore the molecular mechanism of how brassinolide improves the cold resistance of tea plants, "Qiancha 1" was used as the material, and the method of spraying brassinolide on the leaves was adopted to explore its effects on the tea plants under 4 °C low-temperature treatment. The results showed that brassinolide can significantly increase the protective enzyme activity of tea plants under cold stress and reduce cold damage. At the transcriptome level, brassinolide significantly enhanced the expression of key genes involved in calcium signal transduction, Calmodulin (CaM), Calcium-dependent protein kinase (CDPK), calcineurin B-like protein (CBL) and calmodulin-binding transcriptional activators (CAMTA), which then activated the downstream key genes transcriptional regulator CBF1 (CBF1) and transcription factor ICE1 (ICE1) during cold induction. Quantitative real-time PCR (qRT‒PCR) results showed that the expression of these genes was significantly induced after treatment with brassinolide, especially CaM and CBF1. When calcium signalling was inhibited, the upregulated expression of CBF1 and ICE1 disappeared, and when CAMTA was knocked down, the expression of other genes under cold stress was also significantly reduced. The above results indicate that brassinolide combined with the calcium signalling pathway can improve the cold resistance of tea plants. This study provides a new theoretical basis for the study of the cold resistance mechanism of brassinolide.

Exogenous brassinosteroid alleviates calcium deficiency induced tip-burn by regulating calcium transport in Brassica rapa L. ssp. pekinensis.

Mini Chinese cabbage (Brassica rapa L. ssp. Pekinensis) plays an important role in the supply of summer vegetables on the plateau in western China. In recent years, tip-burn has seriously affected the yield, quality and commodity value of mini Chinese cabbage. Calcium (Ca2+) deficiency is a key inducer of tip-burn. As a new type plant hormone, brassinolide (BR) is involved in regulating a variety of biotic and abiotic stresses. To explore the alleviation role of BR in tip-burn caused by Ca2+ deficiency, a hydroponic experiment was conducted to study the relationship between BR and Ca2+ absorption and transport. The results showed that foliar spraying with 0.5 µM BR significantly reduced tip-burn incidence rate and disease index of mini Chinese cabbage caused by Ca2+ deficiency. Moreover, the dynamic monitoring results of tip-burn incidence rate showed that the value reached the highest on the ninth day after treatment. BR promoted the Ca2+ transport from roots to shoots and from outer leaves to inner leaves by increasing the activities of Ca2+-ATPase and H+-ATPase as well as the total ATP content, which provided power for Ca2+ transport. In addition, exogenous BR upregulated the relative expression levels of BrACA4, BrACA11, BrECA1, BrECA3, BrECA4, BrCAX1, BrCAS and BrCRT2, whereas Ca2+ deficiency induced down-regulation. In conclusion, exogenous BR can alleviate the Ca2+-deficiency induced tip-burn of mini Chinese cabbage by promoting the transport and distribution of Ca2+.

Effects of foliar applications of Brassinolide and Selenium on the accumulation of Arsenic and Cadmium in rice grains and an assessment of their health risk.

Arsenic and cadmium pose a potential health risk to human beings via rice grain consumption. In the current study, a pot experiment was conducted to evaluate the effect of Br (5 mM and 20 mM) and Se (1 mM) at rice tillering and filling stages on Cd and As accumulation in rice grain and their health risk indices. The results showed that Br or Se applications at different stages of rice improved the photosynthesis, reduce MDA content in flag leaves by 17.41%-38.65%, increased rice biomass and grain yield by 10.50%-29.94% and 10.50%-36.56%, and enhanced grain N and P uptake by 3.25%-34.90%, and 22.98%-72.05%, respectively. Applications of Br and Se effectively decreased Cd and As concentration in rice grain by 31.74%-86.97% and 16.42%-81.13% respectively. Compared to the individual treatment, combined 20 mM Br and 1 mM Se at the filling stage showed the lowest accumulation of As (0.149 mg·kg-1) and Cd (0.105 mg·kg-1) in grain, and its health risk index was below the acceptable limits (HRI < 1). This implies that application of Br and Se at the filling stage is a promising strategy for the safe production of rice in As and Cd co-contaminated regions.

In this study, foliar applications of Br and Se at the grain filling and tillering stage demonstrate their effect on As and Cd accumulation. The findings showed that Br and Se resulted in the Se concentration in grains reaching the Se-enriched level, and the accumulation of As and Cd was the lowest. Furthermore, the application of Br and Se decreased lipid peroxidation, promoted N and P uptake, and increased the rate of photosynthesis in the rice plants, which resulted in increasing rice growth and grain yield. The HRI of heavy metals was below the acceptable limits after application of Br and Se.

Brassinosteroids Regulate the Water Deficit and Latex Yield of Rubber Trees.

Brassinolide (BR) is an important plant hormone that regulates the growth and development of plants and the formation of yield. The yield and quality of latex from Hevea brasiliensis are regulated by phytohormones. The understanding of gene network regulation mechanism of latex formation in rubber trees is still very limited. In this research, the rubber tree variety CATAS73397 was selected to analyze the relationship between BR, water deficit resistance, and latex yield. The results showed that BR improves the vitality of rubber trees under water deficit by increasing the rate of photosynthesis, reducing the seepage of osmotic regulatory substances, increasing the synthesis of energy substances, and improving the antioxidant system. Furthermore, BR increased the yield and quality of latex by reducing the plugging index and elevating the lutoid bursting index without decreasing mercaptan, sucrose, and inorganic phosphorus. This was confirmed by an increased expression of genes related to latex flow. RNA-seq analysis further indicated that DEG encoded proteins were enriched in the MAPK signaling pathway, plant hormone signal transduction and sucrose metabolism. Phytohormone content displayed significant differences, in that trans-Zeatin, ethylene, salicylic acid, kinetin, and cytokinin were induced by BR, whereas auxin, abscisic acid, and gibberellin were not. In summary, the current research lays a foundation for comprehending the molecular mechanism of latex formation in rubber trees and explores the potential candidate genes involved in natural rubber biosynthesis to provide useful information for further research in relevant areas.

Multi-Omics Analysis Reveals Synergistic Enhancement of Nitrogen Assimilation Efficiency via Coordinated Regulation of Nitrogen and Carbon Metabolism by Co-Application of Brassinolide and Pyraclostrobin in Arabidopsis thaliana.

Improving nitrogen (N) assimilation efficiency without yield penalties is important to sustainable food security. The chemical regulation approach of N assimilation efficiency is still less explored. We previously found that the co-application of brassinolide (BL) and pyraclostrobin (Pyr) synergistically boosted biomass and yield via regulating photosynthesis in Arabidopsis thaliana. However, the synergistic effect of BL and Pyr on N metabolism remains unclear. In this work, we examined the N and protein contents, key N assimilatory enzyme activities, and transcriptomic and metabolomic changes in the four treatments (untreated, BL, Pyr, and BL + Pyr). Our results showed that BL + Pyr treatment synergistically improved N and protein contents by 56.2% and 58.0%, exceeding the effects of individual BL (no increase) or Pyr treatment (36.4% and 36.1%). Besides synergistically increasing the activity of NR (354%), NiR (42%), GS (62%), and GOGAT (62%), the BL + Pyr treatment uniquely coordinated N metabolism, carbon utilization, and photosynthesis at the transcriptional and metabolic levels, outperforming the effects of individual BL or Pyr treatments. These results revealed that BL + Pyr treatments could synergistically improve N assimilation efficiency through improving N assimilatory enzyme activities and coordinated regulation of N and carbon metabolism. The identified genes and metabolites also informed potential targets and agrochemical combinations to enhance N assimilation efficiency.

Assessment of Biological Activity of 28-Homobrassinolide via a Multi-Level Comparative Analysis.

Brassinosteroids (BRs) play vital roles in the plant life cycle and synthetic BRs are widely used to increase crop yield and plant stress tolerance. Among them are 24R-methyl-epibrassinolide (24-EBL) and 24S-ethyl-28-homobrassinolide (28-HBL), which differ from brassinolide (BL, the most active BR) at the C-24 position. Although it is well known that 24-EBL is 10% active as BL, there is no consensus on the bioactivity of 28-HBL. A recent outpouring of research interest in 28-HBL on major crops accompanied with a surge of industrial-scale synthesis that produces mixtures of active (22R,23R)-28-HBL and inactive (22S,23S)-28HBL, demands a standardized assay system capable of analyzing different synthetic "28-HBL" products. In this study, the relative bioactivity of 28-HBL to BL and 24-EBL, including its capacity to induce the well-established BR responses at molecular, biochemical, and physiological levels, was systematically analyzed using the whole seedlings of the wild-type and BR-deficient mutant of Arabidopsis thaliana. These multi-level bioassays consistently showed that 28-HBL exhibits a much stronger bioactivity than 24-EBL and is almost as active as BL in rescuing the short hypocotyl phenotype of the dark-grown det2 mutant. These results are consistent with the previously established structure-activity relationship of BRs, proving that this multi-level whole seedling bioassay system could be used to analyze different batches of industrially produced 28-HBL or other BL analogs to ensure the full potential of BRs in modern agriculture.

Epi-Brassinolide Regulates ZmC4 NADP-ME Expression through the Transcription Factors ZmbHLH157 and ZmNF-YC2.

Maize is a main food and feed crop with great production potential and high economic benefits. Improving its photosynthesis efficiency is crucial for increasing yield. Maize photosynthesis occurs mainly through the C4 pathway, and NADP-ME (NADP-malic enzyme) is a key enzyme in the photosynthetic carbon assimilation pathway of C4 plants. ZmC4-NADP-ME catalyzes the release of CO2 from oxaloacetate into the Calvin cycle in the maize bundle sheath. Brassinosteroid (BL) can improve photosynthesis; however, its molecular mechanism of action remains unclear. In this study, transcriptome sequencing of maize seedlings treated with epi-brassinolide (EBL) showed that differentially expressed genes (DEGs) were significantly enriched in photosynthetic antenna proteins, porphyrin and chlorophyll metabolism, and photosynthesis pathways. The DEGs of C4-NADP-ME and pyruvate phosphate dikinase in the C4 pathway were significantly enriched in EBL treatment. Co-expression analysis showed that the transcription level of ZmNF-YC2 and ZmbHLH157 transcription factors was increased under EBL treatment and moderately positively correlated with ZmC4-NADP-ME. Transient overexpression of protoplasts revealed that ZmNF-YC2 and ZmbHLH157 activate C4-NADP-ME promoters. Further experiments showed ZmNF-YC2 and ZmbHLH157 transcription factor binding sites on the -1616 bp and -1118 bp ZmC4 NADP-ME promoter. ZmNF-YC2 and ZmbHLH157 were screened as candidate transcription factors mediating brassinosteroid hormone regulation of the ZmC4 NADP-ME gene. The results provide a theoretical basis for improving maize yield using BR hormones.

Transcriptome Analysis Reveals Brassinolide Signaling Pathway Control of Foxtail Millet Seedling Starch and Sucrose Metabolism under Freezing Stress, with Implications for Growth and Development.

Low-temperature stress limits the growth and development of foxtail millet. Freezing stress caused by sudden temperature drops, such as late-spring coldness, often occurs in the seedling stage of foxtail millet. However, the ability and coping strategies of foxtail millet to cope with such stress are not clear. In the present study, we analyzed the self-regulatory mechanisms of freezing stress in foxtail millet. We conducted a physiological study on foxtail millet leaves at -4 °C for seven different durations (0, 2, 4, 6, 8, 10, and 12 h). Longer freezing time increased cell-membrane damage, relative conductance, and malondialdehyde content. This led to osmotic stress in the leaves, which triggered an increase in free proline, soluble sugar, and soluble protein contents. The increases in these substances helped to reduce the damage caused by stress. The activities of superoxide dismutase, peroxidase, and catalase increased reactive oxygen species (ROS) content. The optimal time point for the response to freezing stress was 8 h after exposure. The transcriptome analysis of samples held for 8 h at -4 °C revealed 6862 differentially expressed genes (DEGs), among which the majority are implicated in various pathways, including the starch and sucrose metabolic pathways, antioxidant enzyme pathways, brassinolide (BR) signaling pathway, and transcription factors, according to Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment. We investigated possible crosstalk between BR signals and other pathways and found that BR signaling molecules were induced in response to freezing stress. The beta-amylase (BAM) starch hydrolase signal was enhanced by the BR signal, resulting in the accelerated degradation of starch and the formation of sugars, which served as emerging ROS scavengers and osmoregulators to resist freezing stress. In conclusion, crosstalk between BR signal transduction, and both starch and sucrose metabolism under freezing stress provides a new perspective for improving freezing resistance in foxtail millet.

Boron deficiency-induced root growth inhibition is mediated by brassinosteroid signalling regulation in Arabidopsis.

Brassinosteroids (BRs) are pivotal phytohormones involved in the control of root development. Boron (B) is an essential micronutrient for plants, and root growth is rapidly inhibited under B deficiency conditions. However, the mechanisms underlying this inhibition are still unclear. Here, we identified BR-related processes underlying B deficiency at the physiological, genetic, molecular/cell biological and transcriptomic levels and found strong evidence that B deficiency can affect BR biosynthesis and signalling, thereby altering root growth. RNA sequencing analysis revealed strong co-regulation between BR-regulated genes and B deficiency-responsive genes. We found that the BR receptor mutants bri1-119 and bri1-301 were more insensitive to decreased B supply, and the gain-of-function mutants bes1-D and pBZR1-bzr1-D exhibited insensitivity to low-B stress. Under B deficiency conditions, exogenous 24-epibrassinolide rescued the inhibition of root growth, and application of the BR biosynthesis inhibitor brassinazole exacerbated this inhibitory effect. The nuclear-localised signal of BES1 was reduced under low-B conditions compared with B sufficiency conditions. We further found that B deficiency hindered the accumulation of brassinolide to downregulate BR signalling and modulate root elongation, which may occur through a reduction in BR6ox1 and BR6ox2 mRNA levels. Taken together, our results reveal a role of BR signalling in root elongation under B deficiency.

Phloroglucinol Promotes Fucoxanthin Synthesis by Activating the cis-Zeatin and Brassinolide Pathways in Thalassiosira pseudonana.

Phloroglucinol improves shoot formation and somatic embryogenesis in several horticultural and grain crops, but its function in microalgae remains unclear. Here, we found that sufficiently high concentrations of phloroglucinol significantly increased fucoxanthin synthesis, growth, and photosynthetic efficiency in the microalga Thalassiosira pseudonana. These results suggested that the role of phloroglucinol is conserved across higher plants and microalgae. Further analysis showed that, after phloroglucinol treatment, the contents of cis-zeatin and brassinolide in T. pseudonana increased significantly, while the contents of trans-zeatin, N6-isopentenyladenine (iP), auxin, and gibberellin were unaffected. Indeed, functional studies showed that the effects of cis-zeatin and brassinolide in T. pseudonana were similar to those of phloroglucinol. Knockout of key enzyme genes in the cis-zeatin synthesis pathway of T. pseudonana or treatment of T. pseudonana with a brassinolide synthesis inhibitor (brassinazole) significantly reduced growth and fucoxanthin content in T. pseudonana, and phloroglucinol treatment partially alleviated these inhibitory effects. However, phloroglucinol treatment was ineffective when the cis-zeatin and brassinolide pathways were simultaneously inhibited. These results suggested that the cis-zeatin and brassinolide signaling pathways are independent regulators of fucoxanthin synthesis in T. pseudonana and that phloroglucinol affects both pathways. Thus, this study not only characterizes the mechanism by which phloroglucinol promotes fucoxanthin synthesis but also demonstrates the roles of cis-zeatin and brassinolide in T. pseudonana. IMPORTANCE Here, we demonstrate that phloroglucinol, a growth promoter in higher plants, also increases growth and fucoxanthin synthesis in the microalga Thalassiosira pseudonana and therefore may have substantial practical application for industrial fucoxanthin production. Phloroglucinol treatment also induced the synthesis of cis-zeatin and brassinolide in T. pseudonana, and the cis-zeatin and brassinolide signaling pathways were implicated in the phloroglucinol-driven increases in T. pseudonana growth and fucoxanthin synthesis. Thus, our work clarified the molecular mechanism of phloroglucinol promoting the growth and fucoxanthin synthesis of Thalassiosira pseudonana and suggested that cis-zeatin and brassinolide, in addition to phloroglucinol, have potential utility as inducers of increased microalgal fucoxanthin production.

Effects of brassinosteroids on cancer cells: A review.

Brassinolide is a new type of steroidal hormone with strong activities, which is well known as an efficient and low-toxicity plant growth regulator for a long time. Because steroidal hormones have a wide application prospect, brassinosteroids have been gradually explored in pharmacology and animal cells in recent decades. Brassinolide could effectively reverse the resistance of human T lymphoblastoid cell line CCRF-VCR 1000 by inhibiting the effusion of drug transported by P-glycoprotein. Brassinosteroids could also accelerate wound healing by positively eliminating inflammation and stimulating reepithelialization of the reparation stage. The occurrence of cancer is a multistep process mediated by a variety of factors. Until now, cancer has always been one group of the major diseases that threaten human health. Many studies have found that brassinosteroids have attracted a great deal of potential as an anticancer agent in the treatment of cancer cells, and most of them exert anticancer activity by inducing apoptosis in cancer cells. There are few articles on the relationship between brassinosteroids and cancer so far. Accordingly, in this article, we summarized current research about the brassinosteroids and cancers. Through the review, researchers could know more about brassinosteroids which might become a new tool for the treatment of cancer in the future, and not only a plant hormone.

Brassinolide alleviates Fe deficiency-induced stress by regulating the Fe absorption mechanism in Malus hupehensis Rehd.

KEY MESSAGE: Exogenous brassinolide promotes Fe absorption through mechanism I strategy, thus improving the tolerance of Malus hupehensis seedlings to Fe deficiency stress. Iron (Fe) deficiency is a common nutritional disorder that results in decreased yield and poor fruit quality in apple production. As a highly active synthetic analog of brassinosteroids, brassinolide (BL) plays numerous roles in plant responses to abiotic stresses. However, its role in Fe deficiency stress in apple plants has never been reported. Herein, we found that the exogenous application of 0.2 mg L-1 BL could significantly enhance the tolerance of apple seedlings to Fe deficiency stress and result in a low etiolation rate and a high photosynthetic rate. The functional mechanisms of this effect were also explored. We found that first, exogenous BL could improve Fe absorption through the mechanism I strategy. BL induced the activity of H+-ATPase and the expression of MhAHA family genes, resulting in rhizosphere acidification. Moreover, BL could enhance the activity of Fe chelate reductase and absorb Fe through direct binding with the E-box of the MhIRT1 or MhFRO2 promoter via the transcription factors MhBZR1 and MhBZR2. Second, exogenous BL alleviated osmotic stress by increasing the contents of osmolytes (proline, solution proteins, and solution sugar) and scavenged reactive oxygen species by improving the activities of antioxidant enzymes. Lastly, exogenous BL could cooperate with other endogenous plant hormones, such as indole-3-acetic acid, isopentenyl adenosine, and gibberellic acid 4, that respond to Fe deficiency stress indirectly. This work provided a theoretical basis for the application of exogenous BL to alleviate Fe deficiency stress in apple plants.

Metabolomic profiling of brassinolide and abscisic acid in response to high-temperature stress.

KEY MESSAGE: Hormone treatment enhanced the content of osmotic substances under high-temperature conditions. The effect of ABA and BR treated separately is better than treated together. To determine the effect of abscisic acid (ABA), brassinolide (BR) and ABA + BR on grape quality under high-temperature stress, various metabolites were analyzed. Compared with the control (CK), DL-tryptophan, D-raffinose, geniposidic acid, dodecanedioic acid and polyphenols were found to be higher after ABA treatment. After BR treatment, amino acids and poricoic acid B were higher than in CK. And carbohydrates and amino acids were up-regulated after ABA + BR treatment. BR and ABA + BR treatment also induced higher endogenous ABA and epibrassinolide contents. In addition, treated grape had higher soluble solid concentrations and soluble sugar content, and delayed the degradation of middle lamella and microfibrils. Antioxidant and heat shock-related genes were examined, which significantly increased in treated grape. The finding of this study suggested that ABA, BR and ABA + BR are very useful for alleviating high-temperature damage by increasing the accumulation of osmotic adjustment substances, and endogenous hormones content.

Structure modification of nonsteroidal brassinolide-like compound, NSBR1.

Brassinolide (BL) is a possible plant growth regulator in agriculture, but the presence of a steroid skeleton hampers the field application of BL in agriculture because of its high synthetic cost. We discovered NSBR1 as the first nonsteroidal BL-like compound using in silico technology. Searching for more potent BL-like compounds, we modified the structure of NSBR1 with respect to 2 benzene rings and the piperazine ring. The activity of synthesized compounds was measured in Arabidopsis hypocotyl elongation. The propyl group of butyryl moiety of NSBR1 was changed to various alkyl groups, such as straight, branched, and cyclic alkyl chains. Another substituent, F, at the ortho position of the B ring toward the piperazine ring was changed to other substituents. A methyl group was introduced to the piperazine ring. Most of the newly synthesized compounds with the 3,4-(OH)2 group at the A ring significantly elongated the hypocotyl of Arabidopsis.

Exogenous 24-epibrassinolide boosts plant growth under alkaline stress from physiological and transcriptomic perspectives: The case of broomcorn millet (Panicum miliaceum L.).

Land alkalization is an abiotic stress that affects global sustainable agricultural development and the balance of natural ecosystems. In this study, two broomcorn millet cultivars, T289 (alkaline-tolerant) and S223 (alkaline-sensitive), were selected to investigate the response of broomcorn millet to alkaline stress and the role of brassinolide (BR) in alkaline tolerance. Phenotypes, physiologies, and transcriptomes of T289 and S223 plants under only alkaline stress (AS) and alkaline stress with BR (AB) were compared. The results showed that alkaline stress inhibited growth, promoted the accumulation of soluble sugars and malondialdehyde, enhanced electrolyte leakage, and destroyed the integrity of broomcorn millet stomata. In contrast, BR lessened the negative effects of alkaline stress on plants. Transcriptome sequencing analysis showed that relative to control groups (CK, nutrient solution), in AS groups, 21,113 and 12,151 differentially expressed genes (DEGs) were identified in S223 and T289, respectively. Gene Ontology (GO) and the Kyoto Encyclopedia of Genes and Genomes (KEGG) revealed various terms and pathways related to metabolism. Compared to S223, alkaline stress strongly activated the brassinosteroid biosynthesis pathway in T289. Conversely, ARF, TF, and TCH4, associated with cell growth and elongation, were inhibited by alkaline stress in S223. Moreover, alkaline stress induced the activation of the mitogen-activated protein kinase (MAPK) pathway, the abscisic acid signaling pathway that initiates stomatal closure, as well as the starch and sucrose metabolism. The EG and BGL genes, which are associated with cellulose degradation, were notably activated. BR enhanced alkaline tolerance, thereby alleviating the transcriptional responses of the two cultivars. Cultivar T289 is better in alkalized regions. Taken together, these results reveal how broomcorn millet responds to alkaline stress and BR mitigates alkaline stress, thus promoting agriculture in alkalized regions.

Principal Component Analysis to Assess the Changes of Yield and Quality in Pinellia ternata at Different Stages after Brassinolide Treatments.

Brassinolide (BR) is the "sixth class" plant hormone, which plays an important role in various physiological and biochemical processes of plants. The wide variety of functions of Pinellia ternata means that there is huge demand for it and thus it is in short supply. This paper mainly assessed the changes of yield and quality in P. ternata at different stages after BR treatments by principal component analysis, in order to improve the yield and quality of P. ternata and at the same time determine the best harvest time. The results showed that the tuber yield of P. ternata was significantly increased by BR treatments at different stages (except for the 15th day). After the 15th, 45th, 60th, 75th, 90th, and 105th day of treatments, the tuber yield of P. ternata reached peak values at 0.10 (0.65 g), 0.50 (1.97 g), 0.50 (1.98 g), 1.00 (2.37 g), 1.00 (2.84 g), and 2.00 mg/L (3.76 g) BR treatment, respectively. The optimal harvest time was the 75th day after 0.10, 0.50, and 1.00 mg/L BR treatments, which not only significantly improved the yield of P. ternata, but also retained high level of total alkaloids in the tubers (20.89, 5.37, and 13.44%) and bulbils (9.74, 20.42, and 13.62%), high total flavone content in the tubers (17.66, 16.26, and 12.74%) and bulbils (52.63, 12.79, and 38.69%), and high ß-sitosterol content in the tubers (25.26, 16.65, and 0.62%) of P. ternata, compared with the control, respectively.

Transcriptome Analysis Reveals an Essential Role of Exogenous Brassinolide on the Alkaloid Biosynthesis Pathway in Pinellia Ternata.

Pinellia ternata (Thunb.) Druce is a traditional medicinal plant containing a variety of alkaloids, which are important active ingredients. Brassinolide (BR) is a plant hormone that regulates plant response to environmental stress and promotes the accumulation of secondary metabolites in plants. However, the regulatory mechanism of BR-induced alkaloid accumulation in P. ternata is not clear. In this study, we investigated the effects of BR and BR biosynthesis inhibitor (propiconazole, Pcz) treatments on alkaloid biosynthesis in the bulbil of P. ternata. The results showed that total alkaloid content and bulbil yield was enhanced by 90.87% and 29.67% under BR treatment, respectively, compared to the control. We identified 818 (476 up-regulated and 342 down-regulated) and 697 (389 up-regulated and 308 down-regulated) DEGs in the BR-treated and Pcz-treated groups, respectively. Through this annotated data and the Kyoto encyclopedia of genes and genomes (KEGG), the expression patterns of unigenes involved in the ephedrine alkaloid, tropane, piperidine, pyridine alkaloid, indole alkaloid, and isoquinoline alkaloid biosynthesis were observed under BR and Pcz treatments. We identified 11, 8, 2, and 13 unigenes in the ephedrine alkaloid, tropane, piperidine, and pyridine alkaloid, indole alkaloid, and isoquinoline alkaloid biosynthesis, respectively. The expression levels of these unigenes were increased by BR treatment and were decreased by Pcz treatment, compared to the control. The results provided molecular insight into the study of the molecular mechanism of BR-promoted alkaloid biosynthesis.