Effects of Lactone- and Ketone-Brassinosteroids of the 28-Homobrassinolide Series on Barley Plants under Water Deficit.

The aim of this work was to study the ability of 28-homobrassinolide (HBL) and 28-homocastasterone (HCS) to increase the resistance of barley (Hordeum vulgare L.) plants to drought and to alter their endogenous brassinosteroid status. Germinated barley seeds were treated with 0.1 nM HBL or HCS solutions for two hours. A water deficit was created by stopping the watering of 7-day-old plants for the next two weeks. Plants responded to drought through growth inhibition, impaired water status, increased lipid peroxidation, differential effects on antioxidant enzymes, intense proline accumulation, altered expression of genes involved in metabolism, and decreased endogenous contents of hormones (28-homobrassinolide, B-ketones, and B-lactones). Pretreatment of plants with HBL reduced the inhibitory effect of drought on fresh and dry biomass accumulation and relative water content, whereas HCS partially reversed the negative effect of drought on fresh biomass accumulation, reduced the intensity of lipid peroxidation, and increased the osmotic potential. Compared with drought stress alone, pretreatment of plants with HCS or HBL followed by drought increased superoxide dismutase activity sevenfold or threefold and catalase activity (by 36%). The short-term action of HBL and HCS in subsequent drought conditions partially restored the endogenous B-ketone and B-lactone contents. Thus, the steroidal phytohormones HBL and HCS increased barley plant resistance to subsequent drought, showing some specificity of action.

Structurally Different Exogenic Brassinosteroids Protect Plants under Polymetallic Pollution via Structure-Specific Changes in Metabolism and Balance of Cell-Protective Components.

Heavy metals and aluminum are among the most significant abiotic factors that reduce the productivity and quality of crops in acidic and contaminated soils. The protective effects of brassinosteroids containing lactone are relatively well-studied under heavy metal stress, but the effects of brassinosteroids containing ketone are almost unstudied. Moreover, there are almost no data in the literature on the protective role of these hormones under polymetallic stress. The aim of our study was to compare the stress-protective effects of lactone-containing (homobrassinolide) and ketone-containing (homocastasterone) brassinosteroids on the barley plant's resistance to polymetallic stress. Barley plants were grown under hydroponic conditions; brassinosteroids, increased concentrations of heavy metals (Mn, Ni, Cu, Zn, Cd, and Pb), and Al were added to the nutrient medium. It was found that homocastasterone was more effective than homobrassinolide in mitigating the negative effects of stress on plant growth. Both brassinosteroids had no significant effect on the antioxidant system of plants. Both homobrassinolide and homocastron equally reduced the accumulation of toxic metals (except for Cd) in plant biomass. Both hormones improved Mg nutrition of plants treated with metal stress, but the positive effect on the content of photosynthetic pigments was observed only for homocastasterone and not for homobrassinolide. In conclusion, the protective effect of homocastasterone was more prominent compared to homobrassinolide, but the biological mechanisms of this difference remain to be elucidated.

Synthesis of sulfated brassinosteroids.

A number of water soluble sulfates of 24-epibrassinolide including the 2α,3α-disulfate and all possible monosulfates were synthesized. The target compounds were isolated in crystalline form as the corresponding sodium salts. Pyridine-sulfur trioxide complex was used as sulfating agent followed by transformation of the resulting pyridinium salts into the sodium sulfates by treatment with NaOH. The control of the regioselectivity was achieved by an appropriate use of acetyl and benzyl protecting groups.

Effects of 24-epibrassinolide and green light on plastid gene transcription and cytokinin content of barley leaves.

In order to evaluate whether brassinosteroids (BS) and green light regulate the transcription of plastid genes in a cross-talk with cytokinins (CKs), transcription rates of 12 plastid genes (ndhF, rrn23, rpoB, psaA, psaB, rrn16, psbA, psbD, psbK, rbcL, atpB, and trnE/trnY) as well as the accumulation of transcripts of some photoreceptors (PHYA, CRY2, CRY1A, and CRY1B) and signaling (SERK and CAS) genes were followed in detached etiolated barley leaves exposed to darkness, green or white light ±1µm 24-epibrassinolide (EBL). EBL in the dark was shown to up-regulate the transcription of 12 plastid genes, while green light activated 10 genes and the EBL combined with the green light affected the transcription of only two genes (psaB and rpoB). Green light inhibited the expression of photoreceptor genes, except for CRY1A. Under the green light, EBL practically did not affect the expression of CRY1A, CAS and SERK genes, but it reduced the influence of white light on the accumulation of CAS, CRY1A, CRY1B, and SERK gene transcripts. The total content of BS in the dark and under white light remained largely unchanged, while under green light the total content of BRs (brassinolide, castasterone, and 6-deoxocastasterone) and HBRs (28-homobrassinolide, 28-homocastasterone, and 6-deoxo-28-homocastasterone) increased. The EBL-dependent up-regulation of plastome transcription in the dark was accompanied by a significant decrease in CK deactivation by O-glucosylation. However, no significant effect on the content of active CKs was detected. EBL combined with green light moderately increased the contents of trans-zeatin and isopentenyladenine, but had a negative effect on cis-zeatin. The most significant promotive effect of EBL on active CK bases was observed in white light. The data obtained suggest the involvement of CKs in the BS- and light-dependent transcription regulation of plastid genes.

Brassinosteroid-BODIPY conjugates: Design, synthesis, and properties.

Three BS-BODIPY (brassinosteroids-4,4-difluoro-4-bora-3a,4a-diaza-s-indacene) conjugates were synthesized and their fluorescent and immunological properties were investigated. Two of the conjugates, having present all the functional groups characteristic of BS, were shown to be potentially useful as biological probes to study involvement of BS into physiological processes in living cells.

A new ELISA for quantification of brassinosteroids in plants.

Starting from (22R,23R)-2α,3α,22,23,26-pentahydroxy-5α-cholestan-6-one 26-hemisuccinate, conjugates of 28-norcastasterone with horse radish peroxidase and bovine serum albumin were prepared. The latter conjugate was injected into rabbits; produced polyclonal antibodies were used to quantitate 6-keto-brassinosteroids. The newly developed analytical system was used in combination with two other immunoenzymatic assays for brassinosteroids to determine individual compounds of this series. In addition, a direct method of brassinosteroid analysis was proposed. It has the advantage of requiring no sample pretreatment steps such as extraction with organic solvents and chromatography.

Design and studies of novel polyoxysterol-based porphyrin conjugates.

New types of steroid-porphyrin conjugates derived from 20-hydroxyecdysone (20E) and 24-epibrassinolide (EBl) were synthesized. An exceptional regioselectivity in the reaction of both steroids with porphyrin boronic acids was found to give side-chain-conjugated boronic esters as sole products. UV-Vis-, fluorescence and NMR spectroscopy yielded similar data for all the studied compounds confirming the solvent driven supramolecular assembly with formation of J-aggregates. CD measurements of water diluted solutions showed a clear difference between 20E and EBl conjugates. The latter showed a strong supramolecular chirality, whereas 20E J-aggregates did not.

Stereoselective synthesis of 9alpha-hydroxylated ecdysteroids.

Treatment of ecdysteroids with excess of TBAF in THF was shown to proceed with stereoselective oxidation at the 9alpha position of the carbon skeleton. The stereochemistry of the products was confirmed by X-ray analysis. Using this method, 9alpha-hydroxyecdysteroids were obtained in good yield. The results open a route to novel type of natural and modified steroids that are difficult to access otherwise.