| Greenhouse horticulture is an improtant integrative part of modern agriculture;however, product yield and quality are severely reduced by monocropping obstacledue to repeated cultivation of a single agricultural crop in the same land over severalyears. It is worth to note that continuous cropping obstacle becomes a bottleneckproblem needed to be solved. Autotoxicity is a main cause of the continuous croppingobstacle, which exists widely in vegetable crops, especially cucubit crops. Therefore,it is necessary and important to investigate the mechanism of autotoxicity and seekproper solving measures, which plays theoretical and practical roles in the research ofcontinuous cropping obstacle. Cucumber (Cucumis sativus L.) was used as thematerial to investigate the effects of autotoxic agents on the cell cycle,endoreduplication, cell elongation, redox state and storage metabolism duringpostgerminative growth. Meanwhile, the roles of sucrose and ascorbic acid in thealleviation mechanism of autotoxicity in cucumber were studied. The main resultswere as follows:1. The radicle length of cucumber was significantly inhibited by the treatmentswith aqueous root extracts and root exudates of cucumber. The analysis of cellcycle-related gene expression showed that the transcript levels of CDKB, CycA, CycBand CycD3;2 were decreased when treated with aqueous root extracts while there wasno effect on CDKA and CycD3;1 genes. All the six cell cycle-related genes wererepressed at the transcript levels when treated with root exudates. Moreover, greatchanges were induced by aqueous root extracts and root exudates in the relative nucleiDNA content, showing a decreased proportion of 2C and an increased proportion of8C and leading to an increased level of mean C-value (MCV). Thus, the inhibitoryeffects of aqueous root extracts and root exudates on cucumber radicle growth wereprobably related with the downregulation of cell cycle-related genes. The increasedlevel of endoreduplication was possibly due to a protective mechanism on exposure toautotoxic agents.2. Phenylcarboxylic acids, including benzoic and cinnamic derivatives, inhibited cucumber radicle growth in different extents. Among them, cinnamic acid showed themost significant effect. In addition, cinnamic acid showed a dose-dependant inhibitoryeffect on cucumber radicle length. At 6 h after exposure to the 12 kinds ofphenylcarboxylic acids, the transcript levels of the celt cycle-related genes, includingtwo cyclin-dependent kinases (CDKs) and four cyclins were reduced. Among them,transcript of CycB, a marker gene for mitosis showed a remarkable reduction. Thetemporal analysis showed that expression of mitotic genes (CDKB, CycA and CycB)were reduced throughout the whole experiment, while the reduction of the other genes(CDKA, CycD3;1 and CycD3;2) were only observed at earlier hours after treatments.Meanwhile, an enhancement of endoreduplication was observed. Further time courseanalysis indicated that endoreduplication started as early as 6 h after exposure tocinnamic acid. These results provide evidence that exposure to benzoic acids andcinnamic acids could induce rapid and dramatic down-regulation of cell cycle-relatedgenes, especially the mitotic genes, leading to inhibited root growth. Meanwhile, theblock of mitosis caused by phenylcarboxylic acids resulted in an increased level ofendoreduplication.3. The oil mobilization is the major source of sugar, the pivotal substrate ofcarbon and energy metabolism during seed germination and postgerminative stages ofcucumber. We analyzed the effect of cinnamic acid, one of the most importantautotoxins in root exudates of cucumber, on sugar content, respiration and expressionof genes involved in oil mobilization in cucumber. Exposure to cinnamic acid at0.25mM resulted in a significant reduction in growth followed by a decrease in totalsugar, sucrose and hexose contents and this decrease was significantly attenuated by aco-imbibition of 0.5% sucrose. There was a significant decrease in total respirationrate when treated with cinnamic acid and no significant change was observed in plantsafter exposure to co-imbibition of cinnamic acid and sucrose compared with thecontrol. Transcript levels of genes involved in the oil mobilization weredown-regulated in germinated seeds exposed to cinnamic acid and up-regulated bysucrose. Significantly, reduction in respiration and gene transcripts by cinnamic acidwere attenuated by co-imbibition with sucrose. The roles of sucrose as carbon source and signaling molecules were discussed.4. Further study showed that cinnamic acid significantly inhibited the expressionof expansin genes, indicating that cinnamic acid not only blocked the entry intomitosis, but also repressed the cell elongation. In addition, the redox ratios ofAsA/DHA and GSH/GSSG were decreased when treated with cinnamic acid,indicating the reduction/oxidation (redox) state of cell was severely affected.Exogenous addition of sucrose could alleviate the inhibitory effects on cucumberradicle length caused by several benzoic and cinnamic acid derivatives, which wasprobably related to the high transcript levels of cell cycle-related genes and expansins.On another hand, the upregulation of genes involved in the redox transition ofAsA/DHA and GSH/GSSG, as well as the increased redox ratios cause by sucrose waspositive to the plant growth, protecting them from the phytotoxic substances.5. Ascorbic acid at 0.5 mM showed alleviative effect on the inhibition of radiclelength caused by several benzoic and cinnamic acid derivatives. The transcript levelsof cell cycle-related genes, expansins and ascorbic oxidase gene were increased aftertreatment with co-imbibitions of ascorbic acid and cinnamic acid, meanwhile, thetranscript level of peroxidase was decreased. On one hand, ascorbic acid acceleratedthe progression of cell cycle, protecting cells from the oxidative damage; on the otherhand, ascorbic acid could accelerate cell elongation, delay the process of cell wallstiffening. Thus, exogenous addition of ascorbic acid could alleviate the adverse effectof cinnamic acid on the cucumber radicle growth. |
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