Yan Hong Zhou

A role of brassinosteroids in early fruit development in cucumber

Brassinosteroids (BRs) are essential for many biological processes in plants, however, little is known about their roles in early fruit development. To address this, BR levels were manipulated through the application of exogenous BRs (24-epibrassinolide, EBR) or a BR biosynthesis inhibitor (brassinazole, Brz) and their effects on early fruit development, cell division, and expression of cyclin and cyclin-dependent kinases (CDKs) genes were examined in two cucumber cultivars that differ in parthenocarpic capacity. The application of EBR induced parthenocarpic growth accompanied by active cell division in Jinchun No. 4, a cultivar without parthenocarpic capacity, whereas Brz treatment inhibited fruit set and, subsequently, fruit growth in Jinchun No. 2, a cultivar with natural parthenocarpic capacity, and this inhibitory effect could be rescued by the application of EBR. RT-PCR analysis showed both pollination and EBR induced expression of cell cycle-related genes (CycA, CycB, CycD3;1, CycD3;2, and CDKB) after anthesis. cDNA sequences for CsCycD3;1 and CsCycD3;2 were isolated through PCR amplification. Both CsCycD3;1 and CsCycD3;2 transcripts were up-regulated by EBR treatment and pollination but strongly repressed by Brz treatment. Meanwhile, BR6ox1 and SMT transcripts, two genes involved in BR synthesis, exhibited feedback regulation. These results strongly suggest that BRs play an important role during early fruit development in cucumber.

Brassinosteroids promote metabolism of pesticides in cucumber.

Brassinosteroids (BRs) are known to protect crops from the toxicity of herbicides, fungicides and insecticides. It is shown here that application of 24-epibrassinolide (EBR) accelerated metabolism of various pesticides and consequently reduced their residual levels in cucumber ( Cucumis sativus L). Chlorpyrifos, a widely used insecticide, caused significant reductions of net photosynthetic rate (Pn) and quantum yield of PSII (Phi(PSII)) in cucumber leaves. EBR pretreatment alleviated the declines of Pn and Phi(PSII) caused by chlorpyrifos application, and this effect of EBR was associated with reductions of chlorpyrifos residues. To understand how EBR promotes chlorpyrifos metabolism, the effects of EBR on activity and expression of enzymes involved in pesticide metabolism were analyzed. EBR had a positive effect on the activation of glutathione S-transferase (GST), peroxidase (POD), and glutathione reductase (GR) after treatment with chlorpyrifos, although the effect on GR was attenuated at later time points when plants were treated with 1 mM chlorpyrifos. In addition, EBR enhanced the expression of P450 and MRP, which encode P450 monooxygenase and ABC-type transporter, respectively. However, the expression of GST was consistently lower than that of plants treated with only chlorpyrifos. Importantly, the stimulatory effect of EBR on pesticide metabolism was also observed for cypermethrin, chlorothalonil, and carbendazim, which was attributed to the enhanced activity and genes involved in pesticide metabolism. The results suggest that BRs may be promising, environmentally friendly, natural substances suitable for wide application to reduce the risks of human and environment exposure to pesticides.

Hydrogen peroxide is involved in the cold acclimation-induced chilling tolerance of tomato plants

Cold acclimation increases plant tolerance to a more-severe chilling and in this process an accumulation of H(2)O(2) in plants is often observed. To examine the role of H(2)O(2) in cold acclimation in plants, the accumulation of H(2)O(2), antioxidant metabolism, the glutathione redox state, gas exchange and chlorophyll fluorescence were analyzed after cold acclimation at 12/10xa0°C and during the subsequent chilling at 7/4xa0°C in tomato (Solanum lycopersicum) plants. Cold acclimation modestly elevated the levels of H(2)O(2), the gene expression of respiratory burst oxidase homolog 1 (Rboh1) and NADPH oxidase activity, leading to the up-regulation of the expression and activity of antioxidant enzymes. In non-acclimated plants chilling caused a continuous rise in the H(2)O(2) content, an increase in the malondialdehyde (MDA) content and in the oxidized redox state of glutathione, followed by reductions in the CO(2) assimilation rate and the maximum quantum yield of photosystem II (F(v)/F(m)). However, in cold-acclimated plants chilling-induced photoinhibition, membrane peroxidation and reductions in the CO(2) assimilation rate were significantly alleviated. Furthermore, a treatment with an NADPH oxidase inhibitor or H(2)O(2) scavenger before the plants subjected to the cold acclimation abolished the cold acclimation-induced beneficial effects on photosynthesis and antioxidant metabolism, leading to a loss of the cold acclimation-induced tolerance against chilling. These results strongly suggest that the H(2)O(2) generated by NADPH oxidase in the apoplast of plant cells plays a crucial role in cold acclimation-induced chilling tolerance.

Light quality affects incidence of powdery mildew, expression of defence-related genes and associated metabolism in cucumber plants

To determine whether light quality affects the incidence of disease, we exposed cucumber (Cucumis sativus L. cv. Jinyan No. 4) plants at the 4-leaf stage to white and other monochromatic lights and tested the effects on plant response to Sphaerotheca fuliginea, defence-related gene expression and metabolic changes. Exposure to red light resulted in higher levels of H2O2 and salicylic acid (SA), and stronger expression of defence genes such as PR-1 than exposure to white or other monochromatic lights. In comparison, plants grown under purple and blue light had higher activities of phenylalanine ammonia-lyase (PAL) and polyphenoloxidase (PPO) and higher level of flavonoids than plants grown under other lights. Furthermore, plants grown under red light were more resistant whilst plants grown under other monochromatic lights were less resistant to Sphaerotheca fuliginea than plants grown under white light. These results suggest a role of red light in light-enhanced resistance, which correlates with enhanced SA-dependent signaling pathway.

Melatonin enhances thermotolerance by promoting cellular protein protection in tomato plants

Melatonin is a pleiotropic signaling molecule that provides physiological protection against diverse environmental stresses in plants. Nonetheless, the mechanisms for melatonin‐mediated thermotolerance remain largely unknown. Here, we report that endogenous melatonin levels increased with a rise in ambient temperature and that peaked at 40°C. Foliar pretreatment with an optimal dose of melatonin (10 μmol/L) or the overexpression of N‐acetylserotonin methyltransferase (ASMT) gene effectively ameliorated heat‐induced photoinhibition and electrolyte leakage in tomato plants. Both exogenous melatonin treatment and endogenous melatonin manipulation by overexpression of ASMT decreased the levels of insoluble and ubiquitinated proteins, but enhanced the expression of heat‐shock proteins (HSPs) to refold denatured and unfolded proteins under heat stress. Meanwhile, melatonin also induced expression of several ATG genes and formation of autophagosomes to degrade aggregated proteins under the same stress. Proteomic profile analyses revealed that protein aggregates for a large number of biological processes accumulated in wild‐type plants. However, exogenous melatonin treatment or overexpression of ASMT reduced the accumulation of aggregated proteins. Aggregation responsive proteins such as HSP70 and Rubisco activase were preferentially accumulated and ubiquitinated in wild‐type plants under heat stress, while melatonin mitigated heat stress‐induced accumulation and ubiquitination of aggregated proteins. These results suggest that melatonin promotes cellular protein protection through induction of HSPs and autophagy to refold or degrade denatured proteins under heat stress in tomato plants.

HsfA1a upregulates melatonin biosynthesis to confer cadmium tolerance in tomato plants.

Melatonin regulates broad aspects of plant responses to various biotic and abiotic stresses, but the upstream regulation of melatonin biosynthesis by these stresses remains largely unknown. Herein, we demonstrate that transcription factor heat‐shock factor A1a (HsfA1a) conferred cadmium (Cd) tolerance to tomato plants, in part through its positive role in inducing melatonin biosynthesis under Cd stress. Analysis of leaf phenotype, chlorophyll content, and photosynthetic efficiency revealed that silencing of the HsfA1a gene decreased Cd tolerance, whereas its overexpression enhanced plant tolerance to Cd. HsfA1a‐silenced plants exhibited reduced melatonin levels, and HsfA1a overexpression stimulated melatonin accumulation and the expression of the melatonin biosynthetic gene caffeic acid O‐methyltransferase 1 (COMT1) under Cd stress. Both an in vitro electrophoretic mobility shift assay and in vivo chromatin immunoprecipitation coupled with qPCR analysis revealed that HsfA1a binds to the COMT1 gene promoter. Meanwhile, Cd stress induced the expression of heat‐shock proteins (HSPs), which was compromised in HsfA1a‐silenced plants and more robustly induced in HsfA1a‐overexpressing plants under Cd stress. COMT1 silencing reduced HsfA1a‐induced Cd tolerance and melatonin accumulation in HsfA1a‐overexpressing plants. Additionally, the HsfA1a‐induced expression of HSPs was partially compromised in COMT1‐silenced wild‐type or HsfA1a‐overexpressing plants under Cd stress. These results demonstrate that HsfA1a confers Cd tolerance by activating transcription of the COMT1 gene and inducing accumulation of melatonin that partially upregulates expression of HSPs.

Effects of aqueous root extracts and hydrophobic root exudates of cucumber (Cucumis sativus L.) on nuclei DNA content and expression of cell cycle-related genes in cucumber radicles

Allelopathic interactions implicate the inhibition of cell division by allelochemicals. To examine the effects of autotoxic agents on cell cycle and plant growth, germinated cucumber seeds (Cucumis sativus L.) were incubated in solutions containing the aqueous root extracts of cucumber at 1:100, 1:50, 1:25 and 1:10 (w:v), or the hydrophobic root exudates of cucumber at 25, 50 and 100xa0mg·L−1. Aqueous root extracts and hydrophobic root exudates inhibited radicle elongation by 36.47–60.18% and 38.24–62.50%, respectively. The mitosis-specific genes were down-regulated in roots exposed to aqueous root extracts and hydrophobic root exudates. Meanwhile, exposure to either aqueous root extracts or hydrophobic root exudates decreased the proportion of 2C (C-value) and increased the proportion of 8C, leading to an increased mean C-value. We conclude that autotoxic agent-induced inhibition of radicle growth was partly attributed to the down-regulation of cell cycle-related genes and endoreduplication was enhanced under our experimental condition.

24-Epibrassinolide and abscisic acid protect cucumber seedlings from chilling injury

Summary A study investigated the ameliorative effects of the chill-induced inhibition of photosynthesis in cucumber by brassinolides and abscisic acid. Cucumber (Cucumis sativus L) seedlings at the three-leaf stage were sprayed with 24-epibrassinolide (EBS) and abscisic acid (ABA) at different concentrations. After chilling at 8°C, 4°C and 2°C for 3 d, the seedlings were allowed to recover under normal conditions. ABA and EBS pretreatments significantly alleviated chilling injury, increased the maximal photochemical efficiency (Fv/Fm) and net photosynthetic rate (Pn) of the chilled seedlings. Pn and apparent quantum yield of photosynthesis (QY) for seedlings chilled at 4°C declined to 40% and 39% of those for unchilled seedlings respectively, Pn and QY values for those pretreated with ABA and EBS, however, remained almost unchanged compared with those of unchilled seedlings.

Photoinhibition-induced reduction in photosynthesis is alleviated by abscisic acid, cytokinin and brassinosteroid in detached tomato leaves

Detached leaves of tomato (Lycopersicon esculentum Mill.) experienced photoinhibition associated with sharp reductions in net photosynthetic rate (Pn), quantum efficiency of PSII (ΦPSII) and photochemical quenching (qP) even though they were exposed to mild light intensity (400xa0μmolxa0m−2xa0s−1 PPFD) at 28°C. Photoinhibition and the reduction in Pn, ΦPSII and qP, however, were significantly alleviated by 1xa0mgxa0l−1 ABA, 0.1xa0mgxa0l−1N-(2-chloro-4-pyridyl)-N′-phenylurea (CPPU) and 0.01xa0mgxa0l−1 24-epibrassinolide (EBR). Higher concentrations, however, reduced the effects or even exacerbated the occurrence of photoinhibition. Superoxide dismutase and ascorbate peroxidase activity in leaves increased with the increases in ABA concentration within 1–100xa0mgxa0l−1, CPPU concentration within 0.1–10xa0mgxa0l−1 and EBR concentration within 0.01–1.0xa0mgxa0l−1. Catalase and guaiacol peroxidase activity also increased with the increase in EBR concentration but CPPU and ABA treatments at higher concentrations caused a decrease. Malondialdehyde (MDA) content decreased with the increase in CPPU concentration. ABA and EBR, however, decreased MDA concentration only at 1 and 0.01xa0mgxa0l−1, respectively. In conclusion, detached leaves had increased sensitivity to PSII photoinhibition. Photoinhibition-induced decrease in photosynthesis, however, was significantly alleviated by EBR, CPPU and ABA at a proper concentration.

Microbial community responses associated with the development of Fusarium oxysporum f. sp. cucumerinum after 24-epibrassinolide applications to shoots and roots in cucumber

Fusarium wilt, caused by Fusarium oxysporum f. sp. cucumerinum (FO), is one of the major diseases in cucumber (Cucumis sativus) production. Root and foliar applications of 24-epibrassinolide (EBL), an immobile phytohormone with antistress activity, were evaluated for their effects on the incidence of Fusarium wilt and changes in the microbial population and community in roots of cucumber plants. EBL pre-treatment to either roots or shoots significantly reduced disease severity followed by an improved plant growth regardless of the treatment methods applied. EBL applications decreased the Fusarium population on root surfaces and in nutrient solution, but increased the population of fungi and actinobacteria on root surfaces. PCR-DGGE analysis showed that FO-inoculation had significant effects on the bacterial community on root surfaces as expressed by a decreased diversity index and evenness index, but EBL applications alleviated these changes. Moreover, several kinds of decomposing bacteria and growth-promoting bacteria were identified from root surfaces of FO-inoculated plants and EBL-pre-treated plants, respectively. Overall, these results show that the microbial community on root surfaces was affected by a complex interaction between phytohormone-induced resistance and plant pathogens.