Studies On The Function And Mechanism Of Guanine In Stimulating Plant Immunity

Rice is an important food crop in the world,but it is infected by a variety of diseases in the life style,and diseases lead to a significant decline in yield and quality,and poses a major threat to global food production security.Rice bacterial leaf streak and bacterial blight are two major bacterial diseases,rice sheath blight is one of the most destructive fungal diseases in rice.For a long time,chemical fungicides are widely used in agriculture because they directly act on pathogens.However,prolonged excessive use of chemical fungicides can not only lead to increased pathogen resistance,pollution of the ecological and environment,but also threaten human health and even life.Hence,finding green,effective,safe and ecological methods has become a new way to prevent and control pathogens.As a novel biopesticide,plant immune inducers developed by elicitors do not directly kill pathogens,but target plants by promoting growth and enhancing resistance to biotic and abiotic stresses.Previous studies have found that the plant immune inducer Zhi Neng Cong（ZNC）is an extract of the endophytic fungus strain SJ1,which has ultra-high activity and can promote plant growth,and enhance plant disease resistance.The ZNC was isolated,purified and identified by liquid chromatography mass spectrometry,and many nucleoside compounds were found to be active parts of ZNC,guanine being one of them.This study aims to explore the role and mechanism of guanine in enhancing disease resistance,with the main results as follows:（1）To investigate the ability and concentration of guanine in triggering disease resistance in plants,we first found that 100 ng/m L was the optimal concentration of guanine against Pseudomonas syringae pv.tomato DC3000 in Arabidopsis and subsequent experiments were conducted at this concentration as well.（2）To determine whether guanine can elicit Arabidopsis immune responses,hydrogen peroxide production was detected by staining with DAB,and callose deposition was detected by aniline blue staining in this study.It was found that guanine induced the burst of reactive oxygen species and callose deposition in Arabidopsis and rice.In addition,guanine treatment can also promote the phosphorylation of MAPK in Arabidopsis.（3）To explore the molecular mechanism of guanine-enhanced resistance to DC3000 in Arabidopsis,we conducted inoculation experiments.The results revealed that Ca²⁺influx was found to be required for guanine-induced defense response,and guanine improved the ability of Arabidopsis to resist DC3000 through ethylene and jasmonic acid signaling.（4）Further studies revealed that guanine could induce disease resistance in rice.Guanine could significantly reduce the lesion length of rice bacterial leaf streak and bacterial blight.Guanine treatment of rice followed by inoculation with Rhizoctonia solani YWK196 resulted in a smaller lesion area in the greenhouse,guanine treatment reduced the lesion length of leaf sheath by 37%in the field.In addition,guanine could promote reactive oxygen species burst and callose deposition in rice.The plate experiment confirmed that guanine could not directly inhibit the growth of bacteria and fungi,which indicated that guanine is capable of stimulating resistance to a wide range of diseases in rice independent of direct antibacterial effect.（5）Transcriptome analysis of rice leaves after guanine treatment for 0,2 and 24 h showed that 3270 and 1606 genes were up-regulated and 3251 and 794 genes were down-regulated at 2 and 24 h,respectively.Some of the genes that were involved in the positive regulation of plant disease resistance,such as Os WRKY13 and Os WRKY53,were significantly up-regulated,while genes that negatively regulated the defense response,such as Os WRKY76 and Os WRKY45 were significantly down-regulated.The results indicated that guanine is involved in regulating the expression of rice disease resistance-related genes.（6）To further study the molecular mechanism of guanine-induced disease resistance in rice,we analyzed multiple hormone signaling pathways and found that guanine promoted ethylene synthesis and response genes expression,participated in rice sheath blight resistance in an Os ETR2-and Os ETR3-dependent ways.Similarly,the expression of genes involved in the jasmonic acid synthesis and signal transduction was also regulated by guanine,and the jasmonic acid receptor Os COI1b was required for guanine-induced resistance to sheath blight.In summary,guanine could directly induce resistance to pathogens in plants.The application of guanine could trigger the immune response of plants,regulate the expression of disease resistance-related genes,and improve the resistance of plants to pathogens via ethylene and jasmonic acid pathways.