A Drosophila gene, HNF-4(D), was selected by cross-hybridization with a probe to rat HNF-4 (hepatocyte nuclear factor 4), a steroid hormone receptor super-family member that plays an important role in liver-specific gene expression. The Drosophila gene matched the mouse gene in 60 out of 66 amino acids in the zinc finger DNA binding domain and in 140 out of 206 amino acids in the domain that specifies dimerization and ligand binding. HNF-4(D) is expressed in developing Drosophila embryos in mid-gut, fat bodies and malpighian tubules, a striking similarity to its limited expression in the adult intestine, liver and kidney of the mouse. Furthermore, Drosophila mutant that has a chromosome deletion spanning the HNF-4(D) locus fails to develop tissues where HNF-4(D) is expressed during late embryogenesis. These findings together with the earlier realization that the rat hepatocyte nuclear factor 3 (HNF-3) and forkhead, a Drosophila gene required for anterior and posterior gut formation, had virtually the identical DNA binding domain, lead us to speculate that a group of genes that participate in gut formation of invertebrates has survived in evolution to perform similar functions in mammals.