A novel homeobox-containing gene has been identified. Its name, Orthopedia (Otp), exemplifies the homology shared by both the orthodenticle and Antennapedia homeodomains. Otp is highly conserved in evolution. In mouse, Otp is expressed only in restricted domains of the developing forebrain, hindbrain, and spinal cord. In Drosophila, otp first appears at gastrulation in the ectodermal proctodeum and later in the hindgut, anal plate, and along the CNS. Here, we compare the Otp-, Distal-less homeobox 1-(DIx1-), Orthodenticle homolog 1-(Otx1-), Otx2-, and Empty spiracles homolog 2-expressing domains. Our results indicate that Otp is expressed along the CNS both in mouse and Drosophila; Otp could specify regional identities in the development of the forebrain and spinal cord; transcription of Otp and DIx1 takes place in alternating hypothalamic regions reminiscent of a segment-like pattern; and the structural and functional conservation could correspond to a conserved function maintained in evolution.