We isolated and mapped the human homeobox gene EVX1. This gene encodes a protein of 407 amino acid residues containing a homeodomain closely related to the Drosophila even-skipped (eve) segmentation gene of the pair-rule class. EVX1 belongs to a small family of vertebrate eve-related homeobox genes including human EVX1 and EVX2 genes, their murine homologs, Evx 1 and Evx 2, and the frog Xhox-3 gene. We previously reported that EVX2 is localized at the 5' end of the HOX4 locus on chromosome 2. We show here that EVX1 is localized at the 5' end of the HOX1 locus on chromosome 7, 48 kb upstream from the most 5' of the eleven HOX1 genes, namely HOX1J. Both EVX genes are transcribed in an opposite orientation as compared to that of adjacent HOX genes. Human HOX1 and HOX4 complex loci appear to be both closely linked to a homeobox gene of the EVX family.