Expressional And Functional Comparisons Of Two General Odorant Binding Proteins In Agrotis Ipsilon(Hufnagel)(Lepidoptera:Noctuidae)

The black cutworm moth Agrotis ipsilon(Hufnagel)(Lepidoptera: Noctuidae)is ranked as one of most destructive agricultural pests in the world because of its highly omnivory,wide distribution and serious damage.Currently chemical control has been regarded as the primary method for managing A.ipsilon in the agricultural production,However,A.ipsilon has become resistant to plenty of insecticides simultaneously.Therefore,it is crucially important to develop the newly green,efficient and environmentally-friendly methods and strategies to control the damages caused by A.ipsilon.This study systematically and deeply investigated the expression and functional differentiation of General Odorant Binding Proteins(GOBPs)through molecular and cellular biological methods and immunohistochemistry studies,aiming to provide a new method for the effective prevention and control of A.ipsilon.The main results are as follows:1.The expression patterns of GOBPs in different tissues of A.ipsilonThe quantitative real-time PCR(qRT-PCR)was used to measure the AipsGOBP1-2 transcript levels in different tissues.Tissue expression profiles indicated that both of them were antennae-specific and more abundant in the female antennae than in the male antennae.2.Expression level of AipsGOBPs and the effect of moth development and matingThe quantitative real-time PCR(qRT-PCR)was used to measure the AipsGOBPs transcript levels at different developmental stages and before and after mating of both sexes.Temporal expression profiles showed that both AipsGOBP1 and AipsGOBP2 began to express in antennae 3 days prior to adult emergence from pupae and reached their highest expression level 3 and 4 days after adult emergence,respectively.Meanwhile,the qRT-PCR results indicated that mating has opposite effects and increased their expression in the female antennae but reduced in the male antennae.3.The expression patterns of AipsGOBPs in different antennal sensillaIn situ hybridization and immunolocalization demonstrated that both AipsGOBP1 and AipsGOBP2 were expressed and co-localized in sensilla basiconica and sensilla trichodea of both sexes.4.Binding characterization of AipsGOBPs to sex pheromones and plant volatilesThe fluorescence competitive binding assays showed that AipsGOBP2 exhibited a high binding affinity in vitro with the two major sex pheromone components Z7-12:Ac and Z9-14:Ac and the four plant volatiles cis-3-hexen-1-ol,oleic acid,dibutyl phthalate and ?-caryophyllene with Ki values less than 5 ?M.AipsGOBP1,on the other hand,showed medium binding affinities with the five A.ipsilon sex pheromones and six plant volatiles.AipsGOBP2 also showed a broader ligand-binding spectrum and a greater ligand-binding affinity than AipsGOBP1 with the tested aldehyde and alcohol sex pheromones of Lepidoptera species.These results suggest that AipsGOBPs may play a dual role in the binding both sex pheromones and plant volatiles,and that AipsGOBP2 may play a more important role than AipsGOBP1.The significance of this study were: we deeply and systematically investigated the function of the AipsGOBPs and clarified the expression pattern of the AipsGOBPs in the antennal sensilla,and explained that the GOBPs of A.ipsilon may have a dual roles in binding both sex pheromones and host plant volatiles.Our findings also revealed the evolutionary mechanism of insect GOBPs,and,finally,provided a theoretical basis for screening and designing a more effective attractant and repellent than sex pheromone.