The Mechanism Of Action Of Toxic Fluoride On Rice Resistance To Southern Rice Black Streaked Dwarf Disease

In view of the inefficient transmitting southern rice black streaked dwarf virus(SRBSDV)method in the laboratory and the unclear mechanism of action of Dufulin protecting rice form SRBSDV,the following works were carried out in this work.1.In this study,we firstly established a novel method for transmitting SRBSDV in rice using bud cutting method.The transmission rates of SRBSDV in rice were investigated via the polymerase chain reaction(PCR)method using SRBSDV S7-1 and S9-1 as the primers and the results demonstrated that the bud cutting method was an efficient persistent-transmitting method with a higher virus transmission rate(> 80.00%)compared with traditional viral transmission method by its only vector of the white-backed planthopper(WBPH)(53.60%).Meanwhile,the SRBSDV P1,P2,P3,P4,P5-1,P5-2,P6,P8,P9-1,P9-2,and P10 proteins were identified in the viruliferous rice seedlings using a proteomics approach,which demonstrated that SRBSDV was successfully transmitted to rice plants via the bud cutting method.Moreover,the results showed that SRBSDV could be successfully transmitted via bud cutting method and plants infected SRBSDV exhibited the symptoms of were similar to those transmitted by WBPH.These symptoms could show as dwarfism,small spikes,barren grains,abnormally short,dark green leaves with ruffles,aerial rootlets and branching at nodes,and small streaked waxy galls on stems.The use of the bud cutting method to generate a cheap,efficient,reliable supply of SRBSDV-infected rice seedlings should aid the development of disease control strategies.2.Based on the above method for transmitting SRBSDV in rice,we established a model for studying the mechanism of action of Dufulin protecting rice form SRBSDV.In this study,we used label-free quantitative proteomics to explore the mechanism of action of Dufulin protecting rice form SRBSDV and a total of 2733 proteins were identified and quantified from Dufulin treated virus-free and virus-infected rice.Quantitative analysis showed that 101 and 64 proteins of rice were highly expressed responding to three different concentrations of Dufulin treatments.Bioinformatics analysis showed that,compared with virus-free rice control,the common up-regulated proteins response to Dufulin stimulation in virus-free rice treated by three different concentrations of Dufulin showed that the cellular component(BP)categories were mainly involved in response to salicylic acid(SA,20.0%)and systemic acquired resistance(SAR,11.8%).And hydrolase activity(11.1%)and peroxidase(POD)activity(9.6%)were the most enriched and molecular function(MF)terms,while extracellular region(10.1%)and membrane(6.5%)were the most enriched cellular component(CC)terms.Comparison with SRBSDV-infect rice control,the common up-regulated proteins response to Dufulin stimulation in SRBSDV-infect rice pretreated by three different concentrations of Dufulin showed that the BP categories were mainly involved in response to reactive oxygen species(ROS,10.9%)and response to SA(6.7%).The main MF categories were heme binding(11.3%)and POD activity(9.6%).The results demonstrated that Dufulin could enhance the expression level of the proteins related SAR including POD,salicylic acid-binding protein 2(SABP2),and pathogenesis-related proteins(PRs).3.To further reveal the results of proteomics,the genes expression of POD1(prx131),POD2(prx122),SABP2(ID: P0643D11.24-1),PR-1(ID: Os01g0382000),PR-3(Rcht2),PR-5(ID: OSJNBb0065L20.2),and G protein(RGB1)were analysis by quantitative real-time PCR(RT-qPCR).The results showed that the genes expression level of POD,PR-1,PR-3,PR-5,SABP2 and G protein were enhanced by Dufulin and the expression level in virus-free group positively,which was in accordance with proteomic analysis as well.In addition,the enzymatic assays results indicated Dufulin treatments can significantly enhance the POD enzymatic activity.In general,our study firstly showed the evidences that Dufulin targeted POD to enhance the ROS level,followed by up-regulation of SABP2 and PRs to activate rice systemic acquired resistance(SAR).