Juvenile Hormone And Insulin Activate Met- And ERK- Signaling Pathways To Regulate Vg Expression In Locusta Migratoria

The migratory locust(Locusta migrataria,Lm)is an important agricultural insect pest,with a preference on the grain crops.The migratory locust is characterized by its high fecunidity,phase change,and migration,which ofen causes periodical outbreak.Locust is a hemimetabolous insect with panoistic ovaires.In locusts,vitellogenin(Vg)is synthesized in the fat body,transported by the hemolymph,and eventually deposited into oocyte via endocytosis.In many insects,Vg expression is regulated by juvenile hormone(JH).Two mechanisms have been proposed for JH-dependent Vg expression: direct activation by JH through its receptor Methoprene-tolerant(Met)and indirect regulation by transcripton factors downstream of JH and Met.Moreover,accummlateive evidences have show nthat the insulin-like peptide(ILP)is involved in the regulation of Vg expression.However,the molecular mechanisms of Vg expression coordinately regulated by JH and ILP remain unclear.Based on the previous literature reports and our preliminary results,this dissertation study was focused on Vg expression by both JH an ILP,using the migratory locust as the model system.Locust has two Vg genes,VgA and VgB.The full-length cDNA sequences of VgA and VgB were cloned by RACE PCR.Their upstream promoter regions were also cloned into vectors for the dual luciferase reporter assays.The recombinant vectors were transfected into Drosophila S2 cells and induced by JH treatment.Polyclonal anti-Vg,anti-Met and anti-?-Actin antibodies were successfully raised and used for the examination of protein expression profilies.Vg was detectable in the previtellogenic stage,increased in the early vitellogenic phase,reached a peak in the middle of vitellogenic phase,and then declined at late vitellogenic phase.Met RNAi led to substiantial decrease of Vg expression levels.Additional application of methoprene on Met-depleted locust could not restore the expression of Vg,indicating the involvement of Met in the induction Vg expression.Knockdown of ERK also caused the remarkable decrease of VgA and VgB expression levels in the fat body,indicating that ERK plays a crucial role in locust vitellogenesis.Application of methoprene on ERKdepleted locusts was unable to restore Vg expression as well.Interestingly,ERK phoshorlyation(P-ERK)was post adult emergence and reached a peak on day 3,then delined.As the JH titers was very low during 0-3 days post adult emergenc,we speculated that P-ERK activated the expression of Vg.These results suggest that ERK may be involved in the regulation of Vg expression orchestrated by JH and ILP.Similar to ERK,JH induced the expression of VgA and VgB within 30 min.Nevertheless,the expression patterns of VgA and VgB appeared to vary.While the expression levels of VgA reached a peak at 30 min post JH treatment and then delined rapidly,the expression of VgB maintain at high levels.Luciferase reporter assays showed that both JH and 20 E stimulate the expression of VgA and VgB.In conclusion,the results in this study indicate that both JH and ILP induce the expression of Vg.In addition to Met,ERK is likely a key modulator in JH-and ILP-stimulated Vg expression.Thus,locust vitellogenesis and reproduction are coordinately regulated by JH and ILP.