While early studies on juvenile hormone (JH)Juvenile hormone (JH) functions during metamorphosis were mainly conducted using classical models of insect physiology such as the kissing bug Rhodnius prolixus and tobacco hornworm Manduca sexta, the fruit fly Drosophila melanogaster, has also made unique contributions to JH research, as exemplified by the isolation of the first Methoprene-tolerant (Met) mutant and the identification of Krüppel homolog 1 (Kr-h1) as a major mediator of JH action. After a key enzyme in JH biosynthesis was cloned and Met was characterized as the long-sought JH receptor in other insect species, the powerful molecular genetic tools in D. melanogaster have greatly advanced our understanding of JH signaling and its regulation. In contrast, some unique aspects of JH signaling in higher dipteran insects, such as the lack of precocious metamorphosis in JH-deficient conditions and the loss of a conserved epoxidase enzyme, pose a significant challenge for JH research in D. melanogaster. Consequently, many JH biosynthetic enzymes remain to be characterized in flies, which has also limited our understanding of how JH biosynthesis is regulated at the molecular level. In this chapter, we aim to summarize our current understanding of JH signaling in D. melanogaster and propose future research directions.