Study On The Function Of Genes Involved In The Synthesis And Signaling Pathway Of Juvenile Hormone In Chinese White Pine Beetle(Dendroctonus Armandi)

Juvenile hormone(JH)and ecdysone(20-hydroxyecdysone,20E)synergistically regulate larval molting and pupation,stimulate the development and maturation of adult reproductive system,and induce diapause and senescence in insect growth and development,which has become a focus of many researchers in recent years.The effects of juvenile hormone on pheromone synthesis in Scolytidae(Dendroctonus armandi)have been studied,as has the cloning of some genes in the mevalonate pathway of juvenile hormone synthesis and their effects on juvenile hormone,but the effects of key enzyme genes and signaling pathway genes on juvenile hormone synthesis in Scolytidae(D.armandi)have not been reported.In this study,we investigated the sequence characteristics,phylogeny and gene expression of key enzymes of juvenile hormone synthesis,allatostatin gene(AST),ecdysone receptor genes,ecdysone and juvenile hormone signaling pathway genes,as well as the functions of juvenile hormone synthesis and signaling pathway genes in D.armandi.The following are the study’s principal findings.1.The full-length sequence of Da JHAMT gene related to juvenile hormone synthesis in D.armandi was obtained,which has typical JHAMT sequence characteristics and has the highest similarity with JHAMT of Dendroctonus ponderosae.The expression of Da JHAMT was the highest in the larval and pupal stages of D.armandi.In response to the induction of juvenile hormone,the expression of Da JHAMT was dramatically increased in the larval and pupal stages.Da JHAMT gene expression was up-regulated in the larval stage and downregulated in the pupal stage after treatment with 20-hydroxyecdysone.RNA interference of Da JHAMT reduced pupal emergent rate and wing atrophy in emergent adults by inhibiting the expression of several genes upstream of the synthesis pathway of juvenile hormone.It was demonstrated that Da JHAMT is a key gene in the juvenile hormone synthesis pathway of D.armandi.2.The full-length sequence of allatostatin gene(Da AST)was obtained,and Da AST was highly expressed in adults,in heads of larvae and pupae,as well as in the anterior midgut of adults;juvenile hormone and 20-hydroxyecdysone significantly increased Da AST expression in larvae,but decreased Da AST expression in pupae,suggesting that Da AST may inhibit juvenile hormone synthesis in corpora allata(CA)of D.armandi.The RNA interference of Da AST significantly up-regulated the expression of Da JHAMT gene of D.armandi,indicating that the Da AST gene of D.armandi had an inhibitory effect on the synthesis of juvenile hormone.At the same time,the survival rate and emergent rate of pupae of D.armandi pupae were dramatically reduced,while the abnormality rate of emergent adults was significantly increased,indicating that allatostatin was essential for D.armandi to sustain normal growth and development.3.The full-length sequences of Da Ec R,Da E75,Da Hr3 and Da FTZ-F1,the key genes of the ecdysone signaling pathway of D.armandi were obtained.Da Ec R,Da E75,Da Hr3 and Da FTZ-F1 were all highly expressed during the molting stage,indicating that Da Ec R,Da E75,Da Hr3 and Da FTZ-F1 genes of D.armandi are involved in the molting and metamorphosis of D.armandi.Da Ec R,Da E75,Da Hr3 and Da FTZ-F1 genes are involved in larval molting,larval pupation,and pupal transformation into adults in D.armandi,and play significant regulatory roles in these processes.4.The full-length sequences of Da Kr-h1 and Da Br-C genes of D.armandi were obtained,and both Da Kr-h1 and Da Br-C genes were highly expressed in the larval stage,with the highest expression of Da Kr-h1 in the anterior midgut and the highest expression of Da Br-C in the adult reproductive organ.The expression of Da Kr-h1 gene in larvae and pupae was downregulated by the inhibition of 20-hydroxyecdysone,while the expression of Da Br-C in larvae and pupae was up-regulated by the induction of 20-hydroxyecdysone,which indicated that Da Kr-h1 was an early response factor of juvenile hormone and Da Br-C was an early response factor of ecdysone in D.armandi.5.RNA interference of Da Kr-h1 significantly affected the expression of ecdysone signaling pathway genes,causing up-regulation of the expression of early response gene Da Br-C and significantly elevated sheath wing developmental malformation in early emergent adults;interference of Da Kr-h1 gene in larval stage caused early development of pupae and early pupae,while interference of Da Kr-h1 gene in pupal stage caused significantly increased.It can be concluded that Da Kr-h1 and Da Br-C are key genes in the signal transduction of juvenile hormone of D.armandi and play an important role in larval pupation and adult emergence,and Da Kr-h1 of D.armandi regulates the synthesis and transduction of ecdysone by suppressing the expression of Br-C,which provides an important basis and direction for the screening of artificial interference targets of D.armandi development.