The process of segmentation, in which the developing embryo is divided into repetitive structures along its antero-posterior (A-P) axis, as a means of organizing and coordinating the body plan is found in a wide range of organisms. In Drosophila, homoeotic genes are involved in all levels of segmental organization and in determining segment identity. The roles of these genes in segmentation have been found mainly by mutational studies, but also by in situ hybridization, which has shown their domains of expression. In contrast to Drosophila, however, embryonic expression of homoeobox-containing genes in vertebrate organisms has not been found to follow a segmental pattern. Vertebrate segmentation can be clearly seen in the mesodermal somites, but repetitive morphological structures in the central nervous system (neuromeres) have only recently been shown to have developmental significance. Neuromeres in the hindbrain (rhombomeres) have been defined as segmental units by their pattern of nerve formation in the developing chick and by the alternating expression of Krox-20, a gene encoding a zinc-finger DNA-binding protein, in the 9.5-day-old mouse. Here we report that a mouse homoeobox-containing gene, Hox-2.9, is expressed in a segment-specific manner in the developing mouse hindbrain. This expression is in a region which is flanked by the regions of expression of Krox-20, and is precisely contained within a single neuromere, rhombomere 4.