The origin of evolutionary novelty is believed to involve both positive selection and relaxed developmental
constraint. In
flies, the redesign of anterior patterning during embryogenesis is a major developmental innovation and the rapidly evolving Hox gene, bicoid (bcd), plays a critical role. We report evidence for relaxation of selective
constraint acting on bicoid as a result of its maternal pattern of gene expression. Evolutionary theory predicts 2-fold greater sequence diversity for maternal effect genes than for zygotically
expressed genes, because natural selection is only half as effective acting on autosomal genes expressed in one sex as it is on genes expressed in both sexes. We sample an individual from ten populations of Drosophila melanogaster and nine populations of D. simulans for polymorphism in the tandem gene duplicates bcd, which is maternally expressed, and zerknllt (zen), which is zygotically expressed. In both species, we find the ratio of bcd to zen nucleotide diversity to be two or more in the coding regions but one in the noncoding regions, providing the first quantitative support for the theoretical prediction of relaxed selective constraint on maternal-effect genes resulting from sex-limited expression. Our results suggest that the accelerated rate of evolution observed for bcd is owing, at least partly, to variation generated by relaxed selective constraint.