| Riptortus pedestris belongs to the Hemiptera(Coreidae).It is a piercing-sucking phloem feeding insect,which mainly damages legume crops.It mainly damages legume crops.It is one of the main reasons that cause soybeans to drop flowers,pods,dry seeds and empty pods.In recent years,it has attracted the attention of researchers because of its role as one of the causes of “Zhengqing” in Huang-Huai-Hai summer soybean growing area,which resulted in serious loss of soybean yield.Studies have found that piercing-sucking insects secrete saliva to help them probe and forage during feeding.The salivary effectors can help insects regulate the defense response of plants.In recent years,the development of transcriptomics and proteomics has also accelerated the development of insect saliva effectors,but the research on insect effectors has mainly focused on insects such as aphids and planthoppers.There is no research report on the salivary effector of R.pedestris,so the mechanism of how the salivary effector participates in the R.pedestris-plant interaction is still unclear.In this study,we combined transcriptome investigation and aphid salivary gland effector analysis to identify 196 candidate effectors in R.pedestris.Using Agrobacerium tumefaciens infiltration assays,two candidate effectors named RP172 and RP191 were characterized to trigger cell death in N.benthamiana.We studied the role of two genes in the interaction between insects and plants through sequence analysis,location observation,bioassay,and the research on impact of plant defense genes.RP172 is a unique gene of R.pedestris.It has no known functional domains and has a typical secretion signal peptide.It is highly expressed in the salivary glands.These evidences indicate that RP172 can be secreted into plant tissues during feeding.The overexpression of RP172 in N.benthamiana can cause obvious cell death and is accompanied by a strong ROS burst,but it does not affect the resistance of N.benthamiana to plant pathogens such as Phytophthora capsici and Sclerotinia sclerotiorum.Next,we study whether RP172 can cause the expression of PTI and hormone pathway maker genes.The overexpression of RP172 significantly promoted the up-regulated expression of Acre31,PTI5,WRKY7,and WRKY8 related to the PTI immune pathway.RP172 significantly promoted the expression of SA hormone pathway related genes PR1,PR2 B,PRb-1b,and had no significant effect on the expression of JA and ET hormone pathway related maker genes.RP191 has many similarities with RP172.It is a unique gene of R.pedestris,and has a typical secretion signal peptide.It is highly expressed in the salivary glands.Overexpression in N.benthamiana can cause obvious cell death and is accompanied by a strong ROS burst.There are many differences between RP191 and RP172: RP191 has a known functional domains-Stage III sporulation protein AE.The overexpression of RP191 in N.benthamiana significantly promotes the susceptibility of N.benthamiana to the plant pathogens Phytophthora capsici and Sclerotinia sclerotiorum.RP191 only promoted the up-regulation of ACRE31,and had no significant effect on the three genes PTI5,WRKY7,and WRKY8.RP191 significantly promoted the expression of SA hormone pathway related genes PR1,PR2 b,PRb-1b and JA hormone pathway maker genes,and had no significant effect on the expression of ET hormone pathway maker genes.Finally,in order to determine the functional domain of RP191,we designed a series of truncated mutants of RP191 and performed corresponding phenotypic experiments.Although the functional domain of RP191 could not be accurately determined,the scope of the functional domain of RP191 was further reduced.In summary,this study discovered two effectors RP171 and RP191 that were characterized to trigger cell death in N.benthamiana,and preliminarily revealed the regulatory role of the two effectors in plant defense response species.This study is of great significance for us to understand the invasion mechanism of R.pedestris,to cultivate insect-resistant crop species,and to discover new prevention and control strategies for R.pedestris. |
