Insulin Signaling Transactivation Regulates The Egg Diapause Of Migratory Locust,Locusta Migratoria L.(Orthoptera: Acridoidea)

Experiments on adult diapause of Drosophila melanogaster and Culex pipiens,and pupal diapause of flesh fly Sarcophaga crassipalpis provide strong evidence that insulin signaling takes part in the regulatory pathway leading to diapause phenotype.Tyrosine phosphorylation of insulin signaling molecules including insulin receptor(IR)and insulin receptor substrate(IRS)regulates insulin signaling,so protein tyrosine phosphatases(PTPs)and protein tyrosine kinases(PTKs)play a coordinating role in regulating insulin signaling.Besides,reactive oxygen species(ROS)inhibits the activity of PTPs in the process of tyrosine kinase receptors(RTKs)activation and PRX5 is responsible for the scavenging of ROS.Therefore,in order to certify whether there is a diapause regulation pathway from PRX5,ROS,and PTP1B/PTK to insulin signaling and egg diapause,we used RNAi mediated PRX5,PTP1 B and PTK gene silencing and inhibitors of PTP1 B and PTK on female adult locusts to study their influence on insulin signaling and locust diapause.Results showed that PRX5 and PTP1 B negatively regulated insulin signaling and increased diapause rate of locust eggs,but PTK activated insulin signaling and decreased diapause rate.The concrete results are as following:1.After RNAi mediated PRX5 and PTP1 B gene silencing,relative m RNA level of IR,IRS,PI3 K and Akt in insulin signaling were increased sifnificantly under long and short photoperiod.All eggs hatched under long photoperiod,however,diapause rate of offspring eggs decreased significantly under short photoperiod.2.After RNAi mediated PTK gene silencing,relative m RNA level of IR,IRS,PI3 K and Akt in insulin signaling were decreased dramatically under long and short photoperiod.Under long photoperiod all eggs hatched,but diapause rate of locust eggs increased significantly under short photoperiod.3.After treated by PTP1B-IN-1,inhibitor of PTP1 B,the relative m RNA level of genes in insulin signaling pathway and phosphorylation level of IR and IRS all changed regularly.Under long and short photoperiod,m RNA level of IR,IRS,PI3 K and Akt and phosphorylation level of IR and IRS all increased dramatically.Diapause rate remained 0 under long photoperiod but decreased significantly under short photoperiod.With respect to energy materials accumulation,the content of glycogen and trehalose increased significantly and the content of triglyceride,saturated fatty acid and unsaturated fatty acid all decreased significantly under long photoperiod,while the content of trehalose and saturated fatty acid increased and the content of glycogen,triglyceride and unsaturated fatty acid decreased markedly under short photoperiod.4.Injected with genistein,PTK inhibitor,m RNA level of IR,IRS,PI3 K and Akt and phosphorylation level of IR and IRS all decreased under long and short photoperiod.Diapause rate of locust eggs under long photoperiod was 0 all the time but increased significantly under short photoperiod.About energy substances,only glycogen content increased,while the content of trehalose,triglyceride,saturated fatty acid and unsaturated fatty acid all decreased significantly under long photoperiod.Under short photoperiod,trehalose and saturated fatty acid content both decreased and the content of glycogen,triglyceride and unsaturated fatty acid all increased dramatically.5.PRX5 was expressed in Pichia pastoris GS115 and biologically active PRX5 protein was achieved.After we injected PRX5 protein into newly emerged female adult locusts reared in non-diapause and diapause-induced condition,diapause rate of locust eggs increased significantly in diapause-induced condition and all eggs hatched in non-diapause condition.