Molecular Analysis of Juvenile Hormone Action in Heliothis Virescens

Proteins involved in juvenile honnone (JH) action are attractive targets for insect control and their identity remains to be discovered. The long tenn goals of these studies are to understand the molecular basis of JH action and use this information for the development of environmentally friendly pest management methods. An analysis of the promoter of Choristoneura fumiferana JH and 20-hydroxyecdysone (20E) responsive JH esterase (Cjjhe) gene identified a 30 bp JH-response region (JHRE) that is sufficient to support both JH induction and 20E suppression of JH induction. Nuclear proteins isolated from CF-203 (C.fumiferana), L57 (Drosophila melanogaster), HvEls (Heliothis virescens) and Aa23 (Aedes albopictus) cells specifically bind to this JHRE. In all four cell lines, JHRE-regulated reporter gene was induced by JH and 20E suppressed JH induction. Because Heliothine moths are the major crop pests in the US and worldwide and JHRE functions in HvEls cells exactly the same way as it functions in CF-203 cells, we propose to conduct our studies using this JHRE and HvEls cell line developed from H virescens. The specific objective of this proposal is to identify and characterize genes that play key roles in JH action. Screening of a cDNA library using JHRE as a probe and screening assays in yeast will be employed to identify genes that play key roles in JH action. The identified genes will be subjected to functional analysis by studying binding of proteins expressed from the identified cDNAs to JHRE and labeled JH, the effect of RNA interference mediated knock out/knock down of their expression on JH induction of a reporter gene regulated by JHRE, and quantitative reverse transcriptase PCR and in situ hybridization analysis of their mRNA in the tissues of Heliothis virescens. The proposed research is relevant to the goals of the NRI because it includes identification and functional characterization of genes from a major crop pest and addresses the Genomics and Future Food and Fiber Production issue.