Understanding the genetic and molecular basis of hybrid incompatibilities is a long-standing and central problem in evolutionary genetics. Despite decades of studies, we understand little about the genetic basis of hybrid sterility and even more lacking is our understanding of the molecular basis of hybrid dysfunction. The Bogota and USA subspecies of Drosophila pseudoobscura provides a paradigmatic case of the earliest stages of speciation. Previously, we have shown that a single gene, Overdrive (Ovd) is essential for both hybrid sterility and segregation distortion in hybrid F1 males between these populations. Little is known, however, about the normal function of Ovd within species and its molecular role in causing hybrid sterility between species.
To understand the molecular basis of hybrid sterility, we performed a comprehensive cytological characterization of spermatogenesis in D. pseudoobscura hybrid F1 males. Our results show that the primary defect in hybrid males is a failure to compact DNA after the last meiotic division. This failure to properly condense DNA leads to an arrest in sperm development later in spermiogenesis, leading to little to no functional sperm. We further employed CRISPR-Cas9 based methods to GFP tag Ovd at its native locus in D. melanogaster. Our studies on the localization patterns of Ovd in the developing male germline in congruence with the hybrid defect suggests that Ovd has a direct effect on chromatin state during spermatogenesis that leads to hybrid incompatibility between these new subspecies. Together, these findings are providing the first insights into the molecular function of Ovd and highlight the role of chromatin state dynamics in the evolution of reproductive barriers between species.