| Summary: | Because Drosophila do not grow after initiation of metamorphosis, their final body size is determined by larval growth rate and duration of the larval growth phase. Drosophila metamorphosis is triggered by the steroid hormone ecdysone, which is produced in the prothoracic gland (PG). Ecdysone synthesis requires expression of the "Halloween" genes, which encode ecdysone biosynthetic enzymes. Growth rate is regulated by Insulin-like peptides, which are released from the insulin-producing cells (IPCs). Genetic ablation of the IPCs decreases growth rate and delays onset of metamorphosis, suggesting that ecdysone synthesis is induced by insulin signaling. Inhibiting PI3 Kinase (PI3K), the major effector of insulin signaling, in the PG similarly delays metamorphosis as a consequence of decreased ecdysone synthesis and decreased Halloween gene expression. In contrast, activating PI3K in the PG advances the onset of metamorphosis and increases Halloween gene expression. Here I report that increased insulin signaling, accomplished inhibiting the protein kinase A pathway in the IPCs increases insulin signaling and increases growth rate but also advances the onset of metamorphosis by increasing expression of at least one Halloween gene. Ecdysone synthesis is promoted by a second peptide hormone, PTTH, which activates Halloween gene expression via the Torso receptor followed by Ras and Raf in the PG. Null mutations in the transcription factor broad (br ) prevent torso transcription and thus prevent Halloween gene expression and metamorphosis. Here I identify Br as the mechanistic link between PI3K activity and Halloween gene expression. I found that PI3K activity is required for br expression by inhibiting the downstream kinase GSK-3. I provide evidence that three nuclear hormone receptors, βFTZ-F1, HR3 and E75, link GSK-3 activity with br expression: RNAi-mediated βFTZ-F1 or HR3 knockdown, or E75A overexpression, in the PG prevents br expression. I also found that ectopic Torso pathway activation, accomplished by expressing the constitutively active Rafgof , restores Halloween gene transcription to larvae lacking br or βFTZ-F1 , suggesting that these larvae fail to express Halloween genes because they fail to transcribe torso . These studies identify a potential molecular mechanism linking growth rate with competence to respond to the PITH metamorphic signal and thus initiate metamorphosis.
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