The human homeobox-containing genes EN1 and EN2 are closely related to the Drosophila pattern formation gene engrailed (en), which may be important in brain development, as shown by gene expression studies during mouse embryogenesis. Here, we have refined the localization of EN1 to human chromosome 2q13-q21 using a mapping panel of rodent/human cell hybrids containing different regions of chromosome 2 and a lymphoblastoid cell line with an interstitial deletion, del(2)(q21-q23.2). This regional assignment of EN1 increases to 22 the number of currently known genes on human chromosome 2q that have homologs on the proximal region of mouse chromosome 1.