The Deleted in AZoospermia (DAZ) genes encode potential RNAbinding proteins that are expressed exclusively in prenatal and postnatal germ cells and are strong candidates for human fertility factors. Here we report the identification of an additional member of the DAZ gene family, which we have called BOULE. With the identification of this gene, it is clear that the human DAZ gene family contains at least three members: DAZ, a Y-chromosome gene cluster that arose 30 -40 million years ago and whose deletion is linked to infertility in men; DAZL, the ''father'' of DAZ, a gene that maps to human chromosome 3 and has homologs required for both female and male germ cell development in other organisms; and BOULE, a gene that we propose is the ''grandfather'' of DAZ and maps to human chromosome 2. Human and mouse BOULE resemble the invertebrate meiotic regulator Boule, the proposed ortholog of DAZ, in sequence and expression pattern and hence likely perform a similar meiotic function. In contrast, the previously identified human DAZ and DAZL are expressed much earlier than BOULE in prenatal germ stem cells and spermatogonia; DAZL also is expressed in female germ cells. These data suggest that homologs of the DAZ gene family can be grouped into two subfamilies (BOULE and DAZL) and that members of the DAZ family evolved from an ancestral meiotic regulator, Boule, to assume distinct, yet overlapping, functions in germ cell development.

It is generally believed that Drosophila melanogaster has no closely related species with which it can produce the viable and fertile hybrids that are essential for the genetic analysis of speciation. Following the recent report of molecular differentiation between a Zimbabwe, Africa, population and two United States populations, we provide evidence that strong sexual isolation exists between the D. melanogaster population in Zimbabwe and populations of other continents. In the presence of males of their own kind, females from most isofemale lines of Zimbabwe would not mate with males from elsewhere; the reciprocal mating is also significantly reduced, but to a lesser degree. The genes for sexual behaviors are apparently polymorphic in Zimbabwe and postmating reproductive isolation between this and other populations has not yet evolved. Whole chromosome substitutions indicate significant genetic contributions to male mating success by both major autosomes, whereas the X chromosome effect is too weak to measure. In addition, the relative mating success between hybrid and pure line males supports the interpretation of strong female choice. These observations suggest that we are seeing the early stages of speciation in this group and that it is driven by sexual selection. The genetic and molecular tractability of D. melanogaster offers great promise for the detailed analysis of this apparent case of incipient speciation.The difficulties in studying the genetics of speciation can arise from several sources. First, the species in question may have diverged beyond the incipient stage. Many genetic differences between them could have accumulated after speciation had been completed and the information on the population genetic dynamics of speciation may have been lost. Ideally, we would like to observe variation in genes of reproductive isolation that are still in the process of becoming fixed. Polymorphisms of such genes within species would suggest speciation inflagrante delicto (1). The second difficulty arises when the species of interest does not lend itself to extensive and detailed genetic analysis. The conspicuous absence of a species that could hybridize with Drosophila melanogaster to produce fertile progeny is the prime example of the second point. To some degree, all studies of the genetics of postmating reproductive isolation encounter the two problems (2). Genetic analysis of premating sexual isolation also confronts a third difficulty in that mating behaviors are often labile (3-8). Finding a system of sexual isolation associated with robust behavioral phenotypes is thus crucial for genetic studies of premating isolation.Recently, a collection of isofemale lines of D. melanogaster from Zimbabwe, Africa, was reported to show a surprisingly high level of DNA sequence divergence at several nuclear genes compared to flies from North American populations (9). In light of previous observations that flies of this species collected over a wide geographical range are very similar in their nuclear DNA polym...

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Sex-specific traits that lead to the production of dimorphic gametes, sperm in males and eggs in females, are fundamental for sexual reproduction and accordingly widespread among animals. Yet the sex-biased genes that underlie these sex-specific traits are under strong selective pressure, and as a result of adaptive evolution they often become divergent. Indeed out of hundreds of male or female fertility genes identified in diverse organisms, only a very small number of them are implicated specifically in reproduction in more than one lineage. Few genes have exhibited a sex-biased, reproductive-specific requirement beyond a given phylum, raising the question of whether any sex-specific gametogenesis factors could be conserved and whether gametogenesis might have evolved multiple times. Here we describe a metazoan origin of a conserved human reproductive protein, BOULE, and its prevalence from primitive basal metazoans to chordates. We found that BOULE homologs are present in the genomes of representative species of each of the major lineages of metazoans and exhibit reproductive-specific expression in all species examined, with a preponderance of male-biased expression. Examination of Boule evolution within insect and mammalian lineages revealed little evidence for accelerated evolution, unlike most reproductive genes. Instead, purifying selection was the major force behind Boule evolution. Furthermore, loss of function of mammalian Boule resulted in male-specific infertility and a global arrest of sperm development remarkably similar to the phenotype in an insect boule mutation. This work demonstrates the conservation of a reproductive protein throughout eumetazoa, its predominant testis-biased expression in diverse bilaterian species, and conservation of a male gametogenic requirement in mice. This shows an ancient gametogenesis requirement for Boule among Bilateria and supports a model of a common origin of spermatogenesis.