Genomic and insulin-mediated control of metabolic homeostasis by the mosquito ecdysone-induced gene E93

Ecdysone-induced protein 93 (E93) is an adult specifier that governs insect pupal-adult conversion. It affects the reproductive transition in adult <i>Aedes aegypti</i> mosquitoes, the significant vectors of numerous devastating human diseases. Here, we show that E93 is essential for maintaining metabolic homeostasis during the reproductive cycle of mosquitoes. <i>E93</i> deficiency led to insufficient production of insulin-like peptide 3 (ILP3) from insulin-producing cells in the brain, resulting in reduced phosphorylation of protein kinase B (Akt), a key regulator in the insulin signaling pathway. This reduction facilitated the nuclear translocation of FoxO and enhanced the activity of glycogen synthase kinase 3β (GSK3β), which in turn respectively activated the transcription of genes encoding phosphoenolpyruvate carboxykinase (PEPCK) during gluconeogenesis and reduced glycogen synthesis. Further insulin rescue and the luciferase activity assays demonstrated that E93 directly inhibited <i>PEPCK</i> transcription. Ultimately, <i>E93</i>-depleted mosquitoes exhibited systemic metabolic reprogramming, characterized by the dysregulation of carbohydrate, lipid, and amino acid metabolism. Our findings establish that E93 orchestrates metabolic homeostasis by coordinating the insulin signaling cascade and directly regulating <i>PEPCK</i> expression, thus providing an intrinsic connection between endocrine signaling and E93-mediated reproduction in mosquitoes.