The Research Of Arabidopsis Nucleotide-binding And Leucine-rich Repeat Genes RPS2 And RPM1(D505V)in Rice Broad-spectrum Resistance

Rice(Oryza sativa L.)is an important staple food crop that feeds more than half of the world's population.However,its production is threatened by many biotic and abiotic stresses.Various pathogenic microorganisms and insect pests affect the rice growth,resulting in yield decrease or quality decline.Rice blast disease,caused by the hemibiotroph ascomycete pathogen Magnaporthe oryzae,is one of the most destructive diseases of rice.Bacterial blight is another devastating diseases of rice,caused by Xanthomonas oryzae pv.oryzae(Xoo).Brown planthopper(BPH)is a rice-specific herbivore and is one of the most destructive insects of rice.Therefore,it is an urgent issue to develop durable and broad-spectrum of rice cultivars against disease and insects.NB-LRR resistance genes play an important role in the plant immune system.The function and activation mechanism of these genes are not fully understand,which greatly limits the application of these genes.The application of the autoactive NB-LRR genes can circuvement the limits and confer plant broad-spectrum resistance.The autoactive NB-LRR genes have potential application values for plant diseases without known resistance genes.The activity of the Arabidopsis NB-LRR protein RPS2 is inhibited by RIN4.We hypothesize that RPS2 should be autoactive in rice due to no RIN4 expressed in rice.Based on the conserved domain of the NB-LRR protein RPM1,we made an autoactive version of RPM1(D505V).We transformed P_Ubi::AtRPS2 and P_Ubi::AtRPM1(D505V)into the Japonica rice cultivar Nipponbare to determine the autoactivity of the two NB-LRR proteins and the effects on broad-spectrum resistance.We obtained the following results:1.The autoactivity of the two genes in rice were proved by the dwarf growth appearing on the transgenic plants with high expressions of the two genes.Some transgenic plants with low expression of the two genes grew normally.2.AtRPS2 and AtRPM1(D505V)transgenic plants showed incompatible interactions with M.oryzae strains GC1-4,YA2-3,TF11-3,ZX2-6 and HN8,and confers significant resistance to these strains.The transgenic plants displayed early and strong induction of reactive oxygen species(ROS),callose deposition,expression of defense-related genes and generated severe HR at the penetration sites after challenged with these M.oryzae strains,which limited the proliferation and invasion of these strains.The transgenic plants showed compatible interaction with M.oryzae strains HLJ19-1,NC22-3 and Zhong1-GFP.ROS burst was inhibited in transgenic plants when inoculated with these strains,and the transgenic rice cells showed high levels of invasive hyphae.In hydrogen peroxide tolerance test,the compatible strains could tolerant higher hydrogen peroxide concentrations than incompatible strains,indicated that the effect of hydrogen peroxide to incompatible strains was weakened.In addition,overexpression of AtRPS2 and AtRPM1(D505V)also confers resistance to bacterial pathogen Xoo and insect pest BPH in transgenic rice,but did not confers resistance to sheath blight disease and bacterial leaf streak.3.The transcriptome analysis showed that the expressions of some defense-related genes were primed to adapt the transformed autoactive NLRs in the transgenic plants,indicated that the basic defense in transgenic plants was stronger than Nipponbare.At the same time,some growth-related genes,such as nitrogen uptake and transport-related genes in transgenic plants were lower than Nipponbare.Therefore,the transgenic plants may exsit"Fitness-Cost".In summary,our study revealed that interfamily transfer of dicot NLRs to monocot species could be functional and provides new insights for engineering disease and pest insect resistance.