Evolution of the neural sex-determination system in insects: The doublesex homolog in crickets functions as an X-linked masculinizing factor to regulate morphological but not behavioral sexual dimorphism

In animals, sex-specific social behaviors, such as courtship and aggression, are controlled by sex-specific neural circuits. To understand the neural mechanisms of sex-specific social behaviors from a comparative perspective, it is necessary to understand the sex determination mechanisms of neural circuits and their evolution across animal lineages. To address this, I have focused on the evolution of the neural sex determination system in insects. In the fruit fly Drosophila melanogaster , a model insect with powerful genetics, the molecular mechanisms underlying neuronal sex determination are well understood, and two transcription factor genes, fruitless and doublesex ( dsx ), have been identified as terminal differentiators. In contrast, those in direct-developing insects, which diverged from the holometabolous lineage, including Drosophila , about 400 million years ago, remain unclear. My previous studies found that the fruitless homolog is unlikely to be integrated into the neuronal sex determination system in direct-developing insects. In this study, I investigated whether the dsx homolog in direct-developing insects contributes to neuronal sex determination to form neural circuits that regulate adult social behaviors. I identified the dsx homolog in the two-spotted cricket Gryllus bimaculatus ( Gryllus dsx gene) and revealed that it has a unique isoform configuration, and its encoded proteins contain a cryptic DNA-binding DM domain with accumulated amino acid substitutions. A knockout study revealed that the Gryllus dsx gene functions as an X-linked morphological masculinizing factor but does not determine the sexual orientation or behavioral patterns of adult social behavior in crickets. These results suggest that, at least in crickets, an unknown sex determination factor(s) other than fruitless and dsx plays a central role in neuronal sex determination.