A conserved fertilization complex bridges sperm and egg in vertebrates

Fertilization, the basis for sexual reproduction, culminates in the binding and fusion of sperm and egg. Although several proteins are known to be crucial for this process in vertebrates, the molecular mechanisms remain poorly understood. Using an AlphaFold-Multimer screen, we identified the protein Tmem81 as part of a conserved trimeric sperm complex with the essential fertilization factors Izumo1 and Spaca6. We demonstrate that Tmem81 is essential for male fertility in zebrafish and mice. In line with trimer formation, we show that Izumo1, Spaca6, and Tmem81 interact in zebrafish sperm and that the human orthologs interact in vitro. Notably, complex formation creates the binding site for the egg fertilization factor Bouncer in zebrafish. Together, our work presents a comprehensive model for fertilization across vertebrates, where a conserved sperm complex binds to divergent egg proteins—Bouncer in fish and JUNO in mammals—to mediate sperm-egg interaction. [Display omitted] •AlphaFold-Multimer predicts trimer formation of sperm Izumo1, Spaca6, and Tmem81•Tmem81 is essential for male fertility in fish and mice•Trimer proteins interact in zebrafish sperm, and human orthologs interact in vitro•The zebrafish trimer interacts with Bouncer on the egg AlphaFold-Multimer predicts complex formation by three sperm proteins conserved in vertebrates. Their interaction is experimentally confirmed and shown to bridge sperm and egg during fertilization by interacting with evolutionarily unrelated egg proteins.