Molecular Mechanism In Whitefly Responding To Defense Of Tamato Host Plant

Due to their high adaptability to host and environment,the MEAM1 and MED cryptic species of Bemisia tabaci,have invaded China and caused serious damage to agricultural production.When feeding on the plant,whitefly will secrete saliva into plant tissue.Effectors in saliva are “powerful weapons” which could enable whitefly response to plant defenses,are important factors that whitefly could feed on hundreds of plant hosts and invaded successfully.In order to reveal the interaction mechanism of the whitefly-plant,the functional factors in saliva and salivary glands of MEAM1 were screened by salivary gland transcriptome,differential expressed gene profile(DEG)and salivary protein mass spectrometry.In this study,some potential secretory proteins were obtained,the major effector was analyzed.The results were displayed as followed.1.Through transcriptome sequencing and bioinformatics analysis,55,630 unigenes were generated from whitefly primary salivary glands.Among them 581 unigenes were predicted as secretory proteins.81.6% of 581 unigenes were annotated.Functional annotation showed that the main functions of the main salivary glands of B,tabaci were processing,synthesis and secretion.Unigenes in salivary gland are enriched in many biological pathways,such as glucose metabolism,amino acid metabolism,detoxification and metabolism,and calcium signaling.The secretory proteins were involved in the early immune response,the regulation of plant defense signals and the degradation of toxic substances in phloem sap.581 potential secretory proteins which contain the elicitors and effectors are meaningful for the research of whitefly-plant interaction.2.150 peptides were identified in whitefly watery saliva by shotgun mass spectrometry.The peptides were divided into 25 groups,such as maltase,glucose dehydrogenase,ATP enzyme,calmodulin,calcium-transporting ATPase,heat shock protein,myosin,group troponin,tubulin,etc.After function annotation,it was found that some of these proteins were involved in external digestion,inhibition of plant defense responses and detoxification of plant defense substances and other biochemical pathways.3.By comparing the differential gene expression profiles of B.tabaci which without anti defense reaction(CK),with immediate anti defense(feeding plants 2h)and with long term anti defense(feeding plant 24h),2,623 unigenes were up-regulated in B.tabaci with immediate anti defense,3,349 unigenes were up-regulated in B.tabaci with long term anti defense.There are 1,236 unigenes up-regulated in B.tabaci that feeding 24 h plant than 2h.This result showed that anti defense of B.tabaci has time effect.When feeding plants 2h compared with feeding solution,the genes of B.tabaci related to anti defense and nutrients metabolism were up-regulated,especially the calcium signaling pathway is activated.When feeding plants 24 h compared with feeding plants 2h,only the nutrient and toxic metabolic pathway were enriched,which indicated that B.tabaci could respond to plant defense in 2h after its stylet piercing into the plant tissue.Through analysis of differential gene expression profiles,some potential salivary secretory proteins was found that might play a significant role in the regulation of whitefly-plant interaction.4.The complete cDNA sequence of LAC1 was cloned,and then analyzed by PCR,RNAi and biology methods.LAC1 was found expressed in all the whole life cycle and highly expressed in the salivary glands(SG)and midgut.LAC1 transcript level was higher in plant-fed than diet-fed whitefly head(containing SG).And LAC1 was found might existe in watery saliva of B.tabaci.Meanwhile,LAC1's transcription level was higher in the head and midgut when whiteflies feed on JA sprayed-tomato plants than on control plants.Silencing LAC1 decreased the survival rate on plant,bycontrast,had marginal effect on the survival rate of B.tabaci raised on an artificial diet.These results indicated that LAC1 enable whitefly to overcome the chemical defenses of host plant,and might act as an effector in saliva.Overall,saliva plays an important role in feeding and responding to plant defense response of B.tabaci,LAC1 may act as an effector playing an important role in whitefly's anti defense response to tomato.Potential salivary secretory proteins found in this study are meaningful for research on insect-plant interactions.These results supply a rich source of target genes for whitefly control and insect resistant plant breeding.