| The whitefly Bemisia tacbaci(Gennadius)(Hemiptera: Aleyraodidae)is a major agricultural pest worldwide.Begomoviruses(family Geminiviridae)transmitted by whiteflies have posed a great threat to crop production in China.Begomoviruses are transmitted by whiteflies in a persistent-circulative manner.Their coat protein(CP)is currently the only structural protein known to be involved in the specific transmission of virus by whitefly.CP may interact with whitefly proteins in complex ways to facilitate the infection of midgut,salivary glands and other tissues by the viruses,leading to effective virus transmission.In addition,once acquired by whitefly,tomato yellow leaf curl virus(TYLCV)can activate the immune responses of whitefly,such as autophagy and antibacterial peptides,which may inhibit virus infection and accumulation.So far,only a few studies have been carried out on the interactions between whiteflies and begomoviruses,and thus our understanding of the complex interactions between whiteflies and begomoviruses is rather limited.In this study,we used yeast two-hybrid nuclear system to screen whitefly proteins with potential to interact with TYLCV CP,and applied the isobaric tags for relative and absolute quantitation(iTRAQ)technique to analyze the changes of protein expression in whiteflies infected with TYLCV or papaya leaf curl China virus(PaLCuCNV).(1)Screening whitefly proteins with potential to interact with TYLCV CP using yeast two-hybrid nuclear systemWe used yeast two-hybrid nuclear system to explore the interactions between TYLCV CP and proteins of Middle East-Asia Minor 1(MEAM1)whitefly.TYLCV CP was used as a bait protein to screen the cDNA library of whitefly,and 15 whitefly proteins that may interact with TYLCV CP were identified.Gene Ontology(GO)and Kyoto Encyclopedia of Genes and Genomes(KEGG)pathway analysis implicated that these 15 whitefly proteins were of different catogeries including metabolic process,cell motility,signal transduction,immune response,among others.We then selected one of the proteins,the tumorous imaginal discs(Tid)protein for further testing and found that Tid could bind to TYLCV CP in vitro,and the DnaJ_C domain of Tid(301-499aa)may be the binding site for TYLCV CP.TYLCV infection in whitefly upregulated the transcription of Tid,and through the interference of ds Tid or antibody against Tid,we found that Tid was unfavourable to virus acquisition by whitefly.Based on the function of Tid reported in studies of mammals,we suggest that accumulation of Tid may be a immune response of the whityefly to TYLCV infection.(2)Analysis of the effects of virus infection on protein expression of whitefly using iTRAQWe used the iTRAQ technique to investigate and compare responses of the Mediterranean(MED)whitefly to the infection of TYLCV and PaLCuCNV.We conducted the following three combinations of comparison,i.e.,TYLCV-infected vs.un-infected,PaLCuCNV-infected vs.un-infected,and TYLCV-infected vs.PaLCuCNV-infected,and identified 259,395 and 74 differently expressed proteins(DEPs)respectively.Bioinformatic analysis indicated that these proteins are associated with catabolic process,metabolic process,transport,defense response,cell cycle,and receptor.The comparisons of TYLCV-infected vs.un-infected and that of PaLCuCNV-infected vs.un-infected revealed some similar DEPs,among which the laminin subunit alpha,dystroglycan,integrin alpha-PS2 and cuticle proteins may be involved in viral transport,and the putative defense protein 3 and PITH domain-containing protein CG6153 may be involved in anti-viral response.However,the 20 S proteasome subunits associated with regulation of virus degradation and accumulation was found only in the comparision of PaLCuCN-infected vs.uninfected,but not in the TYLCV-infected whiteflies,indicating that this protein may be involved only in the suppression of PaLCuCNV accumulation in MED.In summary,based on the data obtained using the yeast two-hybrid nuclear system and bioinformatic analysis,we proposed a network of TYLCV CP interacting with whitefly proteins,and found that the whitefly protein Tid may be involved in the suppression of TYLCV accumulation in whitefly.In addition,using the iTRAQ analysis,we investigated the differences in protein expression in the whitefly body following virus infection,and found some whitefly proteins that may be involved in virus transport and whitefly anti-viral defence.These findings may facilitate the investigations into the complex interaction between whiteflies and begomoviruses. |
