Induction Of P-gp And Atg8 Expression By Exogenous Substances In Helicoverpa Armigera

Helicoverpa armigera is an important worldwide pest.Chemical control is an important approach in IPM.With the extensive use of insecticides,some studies have shown that Helicoverpa armigera has resistance to insecticides and Bt.A lot of researches focus on the modification of molecular targets and the enhancement of detoxification in pest resistance to insecticides.Though most of molecular targets banding with insecticide are located on membrane or inner of cell,there are few investigations explaining for insecticide how to penetrate cell membrane of insects.P-glycoprotein(P-gp)is a transmembrane glycoprotein,which encoded by MDR1 and belongs to the family of ABCB.Through the consumption of ATP,it can transport toxic substances from intracellular to extracellular.So it can protect the tissue from exogenous or endogenous toxic substances,and also can affect the absorption of drugs.Autophagy is a process by which cells degrade their damaged organelles and macromolecules under the regulation of autophagy related genes.Autophagy is ubiquitous in all cells to maintain the homeostasis of cell synthesis and degradation,and it is one of the most important biological phenomena.The protein encoded by Atg8 is a kind of small ubiquitin like protein,which is necessary for the formation of autophagic membrane.The transcriptome Library of Helicoverpa armigera was obtained by high-throughput sequencing.The expression library of Helicoverpa armigera at different developmental stages was obtained by sequencing of expression profiles and digital gene expression(DGE)libraries were constructed in the present study.Based on bioinformatics analysis,we screened a gene encoding multidrug resistance associated protein(MRP).The full length of cDNA(4054 bp)encoding a putative P-glycoprotein gene from Helicoverpa armigera was cloned.This putative P-gp sequence encoded a protein of 1253 amino acids with a molecular mass of approximately 137 kDa.qPCR analyses demonstrated that the expression of P-gp was significantly higher in 4th instar larvae as compared to other developmental stages.P-gp transcripts were more abundant in the head and fat bodies than in the other tissues.Compared with the control,the expression of P-gp reach the peak at 12 h after the treatment by 2-tridecanone in all tissues.However,the expression of P-gp obviously increased during from 12h to 48h after the treatment with abamectin in all tissues.In the treatment group of deltamethrin,the expression of P-gp was highest in 12 h,and the expression was highest in the midgut in the four tissues.In the LC10 chlorantraniliprole treatment group,P-gp expression was highest in 48h.In tissue induced expression,P-gp expression was relatively high in the midgut and fat bodies.In the LC90 chlorantraniliprole treatment group,the expression of P-gp in 12 h was the highest,and the expression of P-gp in integument was the highest compared to the control.Immunohistochemistry analyses also verified that 2-tridecanone and abamectin can induce the increase of P-gp expression.RNAi of P-gp significantly raised the mortality rate of larvae treated by the 2-tridecanone and the abamectin,as compared to control larvae fed with GFP dsRNA.The Helicoverpa armigera Atg8 sequence encoded a protein of 117 amino acids with a molecular mass of approximately 1.39 kDa.qPCR analyses demonstrated that the expression of Atg8 was significantly higher in adult stages as compared to other developmental stages.Atg8 transcripts were more abundant in the head and fat bodies than in the other tissues.Compared with the control,the expression of Atg8 reached the peak at 24 h after treatment with 2-tridecanone in all tissues.However,after treatment with abamectin,the expression of Atg8 increased continually,and was highest at 48 h in all tissues.In the treatment group of deltamethrin,the expression of Atg8 in the head,fat bodies and midgut reached peak at 24 h,but there was no significant difference in integument compared to the control.In the LC10 chlorantraniliprole treatment group,the expression of Atg8 peaked at 24 h after treatment,and the expression was higher in the integument and midgut.In the LC90 chlorantraniliprole treatment group,the expression of Atg8 reached peak at 24 h,and the highest expression was in fat bodies.RNAi of Atg8 significantly raised the mortality rate of larvae treated by the 2-tridecanone and the abamectin,as compared to control larvae fed with dsRNA.In this paper,we studied the expression of P-gp and Atg8 in Helicoverpa armigera,which were induced by different exogenous toxic substances.The results showed that P-gp and Atg8 played an important role in the drug resistance to these exogenous toxic substances.This study illustrated the resistance mechanisms of insects from a new perspective,and provided new ideas for revealing the mechanism of pest resistance.