The first step of the purine de novo synthesis pathway is catalyzed by amidophosphoribosyltransferase (E.C.126.96.36.199) which is encoded by two Prat genes in D. melanogaster, Prat and Prat2. Prat is a retrogene duplication of Prat2, where each gene has a distinct expression pattern. Prat transcription is restricted to proliferating tissues such as imaginal discs and the female germ line. Three conserved putative DNA replication-related element binding factor (DREF) sites lie upstream of the Prat coding region. These elements are upstream of many genes important in cell proliferation. We have found that DREF binds directly upstream of Prat and that the DRE sites associated with its activity are necessary for Prat expression; furthermore, we have determined that a second cis-acting element is present upstream of the Prat gene. Finally, the genes Distal-less, Mi-2 and dMyc, which influence Dref activity, do not appear to affect Prat transcription.