Study On The Pathway And Regulation Mechanism Of The Cadmium Excretion In Tall Fescue Leaves

Heavy metal contamination,especially Cd contamination threaten the ecological environment and human health.In China,a large area of the soil was seriously contaminated by Cd,which directly threatened the safety of agricultural products.Phytoremediation is widely used in Cd contaminated soil for its low-cost and high efficiency.Tall fescue(Festuca arundinacea)is a potential material for phytoremediation based on its high tolerance to Cd.In this study,we found that the leaves of tall fescue showed the function of Cd excretion,which could prevent the plants from being poisoned by excessive accumulation of Cd;based on that,the pathway and physiological characteristics of Cd excretion in the leaves of tall fescue were systematically explored;the regulated effects of different plant growth regulators and Zn on the translocation and excretion of Cd were analyzed;RNA-Seq was used to study the transcriptional regulation of tall fescue with Cd induction,and the detoxication mechanisms of Cd in tall fescue was comprehensively elaborated,providing a theoretical basis for phytoremediation technology of tall fescue based on leaf Cd excretion.The main results of this study are as follows:The time course of Cd excretion in leaves of tall fescue was observed by CLSM(confocal laser scanning microscope)with Cd fluorescence.Within 6-24 hours of Cd treatment,excreted Cd gradually spread down from the tip to the edge of the leaves then covered the whole leaf surface in72 hours after Cd treatment.By means of SEM-EDS(scanning electron microscope-energy dispersive X-ray spectrometric microanalysis)analysis,the structure of hydathode in the tip of tall fescue leaves was found,and the weight percentage of Cd in the crystal on the leaf surface was 5.76%.At the 5th,10th and 15th day of Cd treatment,Cd²⁺was detected in the guttation of the first and second upper leaves.Thus,it can be proved that Cd was excreted in the following way:absorption of Cd by the roots—translocation to the shoots via xylem sap—excretion of Cd by the hydathodes in the leaf tip—attachment of Cd on the leaf surface as the form of crystals.The differences in Cd absorption,accumulation,translocation and excretion capacity of 7 tall fescue varieties of with Cd stress were compared by measuring the growth,photosynthetic parameters and Cd concentrations.The results showed that among the 7 tall fescue varieties,the photosynthetic parameters of jaguar 4G were significantly higher than those of other varieties under Cd treatment,and its decreasing amplitude of P_n,T_r and G_s was the lowest after 21 days of treatment.The Cd excretion ability of tall fescue leaves was significantly different among the 7varieties.Cd concentrations in excretions of elite(blue)and jaguar 4G were 98.63 mg/kg and 80.21 mg/kg,respectively,significantly higher than those of the other 5 varieties.Under Cd stress,jaguar 4G could maintain strong photosynthetic capacity,which can ensure its normal growth,and its strong Cd excretion capacities as well as large biomass are suitable for phytoremediation materials based on leaf Cd excretion mechanism.The effects of Cd stress concentrations,time and leaf position on Cd uptake,translocation,accumulation and excretion were analyzed by measuring the concentrations of Cd and other ions in different parts of tall fescue.With the increase of Cd treatment concentration,Cd concentrations in excretion and shoots both showed an increasing trend,and the excretion factor reached the highest at 75?M Cd treatment.With the increase of Cd treatment time,Cd concentrations in excretion and Cd excretion factor increased,while Cd concentration in shoots first increased and then decreased,and reached the highest at the 28th day.Moreover,the concentrations of Cd in shoots showed significant positive correlation with that in excretion.Cd concentrations in excretion,shoots and Cd excretion factor increased with the aging degree of leaves,indicating that older leaves and dead leaves could accumulate and excrete more Cd.Compared with Zn,Ca,Mg and K,Cd has the lowest translocation factor and the highest excretion factor,so there may be a biased excretion mechanism.5.2%of the Cd in roots was translocated to the shoots and 13.6%of the Cd in shoots was excreted out of the plant through the hydathodes.The effects of 6 plant growth regulators(IAA,SA,6-BA,GA₃,ETH and ABA)on the growth,photosynthetic characteristics,Cd uptake,translocation,accumulation and excretion of tall fescue were investigated by foliar spraying.The results showed that the 6 plant growth regulators had no significant effect on the growth of tall fescue,but ETH and SA could alleviate the inhibition of photosynthesis and transpiration caused by Cd stress,which improves the physiological function of tall fescue under Cd stress.SA and ETH could facilitate the Cd uptake and accumulation of roots and shoots,while GA₃ could facilitate the Cd translocation,so that the ability of Cd accumulation and excretion as well as the efficiency of phytoremediation were significantly enhanced.The regulation of Zn on Cd uptake,accumulation,translocation and excretion in tall fescue was studied by analyzing the interaction between Zn/Cd in roots and shoots.The addition of the mineral element Zn to the nutrient solution inhibited the Cd uptake of roots,but promoted the Cd translocation from roots to shoots,as well as the Cd accumulation and excretion in leaves,while the addition of Cd inhibited the uptake,translocation,accumulation and excretion of Zn.The interaction patterns of Zn and Cd in tall fescue were as follows:they antagonized each other in the roots,while Cd antagonized Zn and Zn synergized Cd in the shoots.The concentrations of Zn,Ca,Mg and K in xylem sap increased because of the addition of Zn,and translocation and accumulation of these nutrients were promoted to ensure the normal growth of tall fescue under the stress of Cd.Meanwhile,it can also promote Cd translocation and excretion,which is an effective way to improve the efficiency of phytoremediation.Illumina Hiseq high-throughput sequencing platform was used to sequence 12 c DNA libraries in the leaves and roots of tall fescue with or without Cd stress,and the response of tall fescue to Cd stress was studied at the transcriptional level.A total of 100,285 Unigene were obtained,including 9292 differentially expressed genes,of which 2180 in leaves,and 7112 in roots.According to the enrichment analysis of differentially expressed genes GO function,all the Unigene and DEGs were divided into three categories,respectively for biological process,cell component and molecular function.51 secondary functions were further divided,among them,the function of detoxification,transporter activity,nucleic acid binding transcription factor activity,antioxidant activity,nutrient reservoir activity and so on may related to heavy metals detoxification,such as detoxification,transporter activity,nucleic acid binding transcription factor activity,antioxidant activity,nutrient reservoir activity and so on.In the enrichment analysis of differentially expressed genes KEGG pathway,we found that KEGG pathway associated with Glutathione metabolism was significantly enriched in the leaves of tall fescue with Cd treatment.After Cd treatment,multiple differentially expressed genes related to cell wall metabolism,antioxidant system,plant hormone signal transduction,translocation of metal ions and transcription factor family were found.These genes combined function on Cd uptake,accumulation,translocation,excretion and detoxification in tall fescue.Through the functional analysis of various DEGs,the molecular mechanisms of Cd uptake,accumulation,translocation and excretion in tall fescue were further understood,and the detoxification pathway was further clarificated.These results can be used to regulate the Cd translocation and excretion so that to improve the efficiency of phytoremediation.