The steroid hormone ecdysone plays a key role in insect development. In Drosophila, ecdysone pulses trigger all major developmental transitions including hatching, molting and puparium formation. While the downstream actions of ecdysone have been well studied, we only have a limited understanding of how ecdysone production is regulated so that distinct pulses are formed during development. We recently started to examine the relationship between heme biosynthesis and ecdysone production, because the majority of steroidogenic enzymes, the cytochrome P450s, require heme as a cofactor. From a genome-wide RNAi screen that we had conducted in collaboration with Mike O’Connor’s and Kim Rewitz’s labs, we carried out a secondary screen aimed at identifying genes involved in the regulation of heme production. This screen was limited to the prothoracic gland (PG), the principal source of larval ecdysteroids, and led to the identification of the 34 candidate genes. PG-specific RNAi against these genes caused larval or pupal arrest and/or significant delays in development. Importantly, all RNAi lines displayed enlarged ring glands, 20 of which exhibited strong red autofluorescence in the PG, consistent with fluorescent heme precursor accumulation in this tissue. Here, we present preliminary findings for two candidate genes, Su(Var)2-10 and AGBE, and how they relate to heme and ecdysone regulation.