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.
While sexual reproduction is widespread among animals, it remains enigmatic to what extent sexual reproduction is conserved and when sex-specific gametogenesis (spermatogenesis and oogenesis) originated in animals. Here we demonstrate the presence of the reproductive-specific protein Boule throughout bilaterally-symmetric animals (Bilateria) and the conservation of its male reproductive function in mice. Examination of Boule evolution in insect and mammalian lineages, representing the Protostome and Deuterostome clades of bilateral animals, failed to detect any evidence for accelerated evolution. Instead, purifying selection is the major force behind Boule evolution. Further investigation of Boule homologs among Deuterostome species revealed reproduction-specific expression, with a strong prevalence of testis-biased expression. We further determined the function of a deuterostomian Boule homolog by inactivating Boule in mice (a representative mammal, a class of Deuterostomes). Like its counterpart in Drosophila (a representative of the opposing Protostome clade), mouse Boule is also required only for male reproduction. Loss of mouse Boule prevents sperm production, resulting in a global arrest of spermatogenesis in remarkable similarity to that of Drosophila boule mutants. Our findings are consistent with a common origin for male gametogenesis among metazoans and reveal the high conservation of a reproduction-specific protein among bilaterian animals.