Identification of the juvenile hormone receptor using its interference with ecdysone signaling II

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The juvenile hormones(JHs) regulate a wide aspect of insect development and reproduction in combination with another hormone, ecdysteroid(Ec). While the knowledge on Ec action at molecular level now constitutes a large body, the molecular mode of JH action is still poorly understood. In this project the author performed the researches below using Anopheles mosquitoes and Drosophila. Based on some literatures that describe the interferences by JH with Ec signaling, the author selected several candidate transcription factors for JH receptors, Among them, the author cloned an arylhydrocarbon receptor homologue(Ahr) from the mosquito, Anopheles Stephensi. When transfected into Drosophila S2 cells, Ahr activated the transcription from a model reporter gene bearing dioxin response elements(XRE) in response to JHs. Further studies, however, revealed that Ahr does not bind directly to the XRE, suggesting the observed JH-dependent transactivation by Ahr is not a direct action. During the course in search for transcription factors responsible for JH responses in the S2 cells, the author cloned the Drosophila Methoprene-tolerant(Met) gene, and performed functional analysis on its product, MET : MET bound to JH III with a Kd of 5.3 nM ; MET fused to yeast GAL4 DNA binding domain exerted JH-dependent regulation on a reporter gene having QAL4 UAS in the transfected S2 cells ; localization of MET was nuclear irrespective of the presence or absence of JH. These results suggest that MET might function as a JH-dependent transcription factor, namely the JH receptor.