Abstract
Juvenile hormone (JH) regulates a wide variety of biological activities in holometabolous insects, ranging from vitellogenesis and caste determination in adults to the timing of metamorphosis in larvae. The mechanism of JH signaling in such a diverse array of processes remains either unknown or contentious. We previously found that the nuclear receptor gene E75A is activated in S2 cells as a primary response to JH. Here, by expressing an intracellular form of JH esterase, we demonstrate that JH must enter the cell in order to activate E75A. To find intracellular receptors involved in the JH response, we performed an RNAi screen against nuclear receptor genes expressed in this cell line and identified the orphan receptor FTZ-F1. Removal of FTZ-F1 prevents JH activation of E75A, whereas overexpression enhances activation, implicating FTZ-F1 as a critical component of the JH response. FTZ-F1 is bound in vivo to multiple enhancers upstream of E75A, suggesting that it participates in direct JH-mediated gene activation. To better define the role of FTZ-F1 in JH signaling, we investigated interactions with candidate JH receptors and found that the bHLH-PAS proteins MET and GCE both interact with FTZ-F1 and can activate transcription through the FTZ-F1 response element. Removal of endogenous GCE, but not MET, prevents JH activation of E75A. We propose that FTZ-F1 functions as a competence factor by loading JH signaling components to the promoter, thus facilitating the direct regulation of E75A gene expression by JH.
Highlights
Juvenile hormone (JH) is a critical insect hormone, but its mechanism of action is contentious
We found that in S2 cells, JH can activate the E75A nuclear receptor gene [37], which plays a critical role during larval development and at the onset of metamorphosis [38]
When we tested RNA samples from the same cells in Northern blot hybridization, we found a characteristic transient profile of E75A induction in control cells; E75A transcripts appeared within 0.5 h after JH administration, and their abundance peaked at 1 h and declined after 1.5 h (Fig. 1B)
Summary
JH is a critical insect hormone, but its mechanism of action is contentious. Results: JH activates E75A through an intracellular pathway utilizing FTZ-F1 and GCE, which form a transcriptionally active heterodimer. Several studies suggest that some actions of JH are mediated by plasma membrane receptors that activate the PKC signaling pathway, as is the case in Drosophila males, where JH stimulates protein synthesis in the accessory glands [9] Another role for PKC in regulating gene expression was proposed in studies of JH signaling in L. migratoria and Choristoneura fumiferana [25, 26]. PKC-mediated phosphorylation reduced the binding of corresponding transcription factors to putative JH response elements, resulting in suppression of JH action Another set of studies provides strong evidence that JH modulates gene expression by acting through intracellular receptors, essentially following the paradigm established by steroid hormones, such as ecdysone. FTZ-F1 directly binds to the regulatory regions of E75A and could function as a competence factor to facilitate the direct regulation of gene expression by JH
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