The Bar mutation B of Drosophila melanogaster and optic morphology mutation Om(JD) of Drosophila ananassae result in suppression of ommatidium differentiation at the anterior portion of the eye. Examination was made to determine the genes responsible for these mutations. Both loci were found to share in common a different type of homeobox gene, which we call "BarH1." Polypeptides encoded by D. melanogaster and D. ananassae BarHl genes consist of 543 and 604 amino acids, respectively, with homeodomains identical in sequence except for one amino acid substitution. A unique feature of these homeodomains is that the phenylalanine residue in helix 3, conserved in all metazoan homeodomains so far examined, is replaced by a tyrosine residue. By Northern blotting, considerably more BarH1 RNA was detected in the Bar mutant than in wild type. P element-mediated transformation showed Bar-like eye malformation to be induced by transient overexpression of the BarH1 gene in the late thirdinstar larvae. Somatic recombination analysis indicated normal gene functions ofthe Bar region, including the BarH1 gene, to be required for normal eye morphogenesis.