Maleness-on-the-Y (MoY) orchestrates male sex determination in major agricultural fruit fly pests

In insects, rapidly evolving primary sex-determining signals are transduced by a conserved regulatory module producing sex-specific proteins that direct sexual differentiation1-4. In the agricultural pest Ceratitis capitata (medfly), a Y-linked Maleness factor (M) is thought to repress the autoregulatory splicing of transformer (Cctra), which is required in XX individuals to establish and maintain female sex determination5,6. Despite the sequencing of the medfly genome, its M factor has remained elusive7. Here, we report the identification of a Y-linked gene, Maleness-on the-Y (MoY), and show that it encodes a small and novel protein which is both necessary and sufficient for medfly male sex determination. CRISPR-induced MoY mutants can develop into fertile XY females and expression of MoY in XX individuals can produce fertile males. indicating that, irrespective of its karyotype, sexual fate of the germline follows that of the soma. In contrast to the molecular diversity of the Y-linked M factors isolated in related or within the same dipteran species8-11, we found Y-linked MoY orthologues in seven other Tephritid species spanning ~111 millions of years of evolution, and confirmed their male determining function in the olive fly (Bactrocera oleae) and the oriental fruit fly (Bactrocera dorsalis). Interestingly, in contrast to M factors of other dipteran species, MoY ectopic embryonic expression fully masculinized XX individuals. MoY evolutionary conservation in a large number of important agricultural pests12 widely distributed over four continents, will facilitate the development of transferable genetic control strategies in these species, for example sterile male release or sex-ratio-distorting gene drives.