Beginning of life: molecular "lock-and-key" of fertilisation found
Genetic studies have identified many proteins that contribute to the first contact between egg and sperm. However, no direct evidence has yet demonstrated how these factors bind or form complexes to carry out their function as a “lock-and-key” mechanism. Now, Andrea Pauli’s lab at the IMP and collaborators combined Artificial Intelligence-driven structural predictions with direct experimental evidence to reveal the formation of a fertilisation complex. Their findings, drawn from studies in zebrafish, mice, and human cells, are published in the journal Cell.
Fertilisation marks the first step in the development of an embryo. It begins with the sperm’s journey to the egg, guided by chemical signals. Upon arrival, the sperm binds to the egg’s surface through specific protein interactions. This binding prepares their membranes to fuse, merging their genetic material to form a zygote—a single cell that will eventually grow into a new life.
Despite the advances in understanding these initial steps, the exact mechanisms that allow the sperm and egg to make contact and ultimately fuse remained a mystery. Unlike most cells in the body, which avoid fusion with other cells and maintain their distinct identities, the sperm and egg are specialised for fusion. This process involves a regulated sequence of molecular events that are not yet understood.
Gallery: "kiss of life" of egg and sperm
Over the past 20 years, many proteins have been identified as essential for the interaction between mammalian sperm and egg. However, only two—Izumo1, found on the surface of the sperm, and Juno, located on the egg’s membrane—have been confirmed to directly bind to each other to facilitate fertilisation.
Using the latest advancements of the Artificial Intelligence (AI) tool AlphaFold, Andrea Pauli’s lab at the IMP and international collaborators now identified a new protein complex that facilitates the first molecular connection between sperm and egg and demonstrated its function in living organisms. The findings, published in the journal Cell, reveal that a fundamental lock-and-key mechanism crucial for fertilisation is shared across vertebrates.
AI unlocks foundations of fertilisation
The fusion of sperm and egg is a highly selective, one-time event that will kickstart the development of a whole new organism. This process relies on a specialised molecular machinery unique to these cell types. Over the years, genetic screens have helped scientists identify several proteins involved.
Researchers turned to AI to go beyond a list of genes important for fertilisation, aiming instead to reveal how these elements function and interact at the molecular level. They used ‘AlphaFold Multimer’, an advanced software that extends the original AlphaFold technology—which predicts individual protein structures based on their sequences—to forecast how different proteins interact with each other and form complexes.
Focusing their initial analysis on proteins known to be found on the surface of sperm, the team employed the AI tool to identify potential additional players in fertilisation. “We assembled a list of proteins predicted to be at the sperm’s membrane and performed a bioinformatic screen including thousands of predictions using AlphaFold,” explains Victoria Deneke, postdoc in the Pauli lab and co-first author of the study. “AlphaFold predicted which proteins might interact with each other and suggested promising candidates for further testing.”
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AcceptThrough this initial screen, they discovered that two previously known fertility-related proteins on the sperm’s surface—Izumo1 and Spaca6—not only interacted with each other, but also with a third, previously unknown factor: Tmem81.“We were surprised to discover a new protein that had never been characterised before,” explains Andreas Blaha, co-first author and a student in the Vienna BioCenter PhD Program. “What really excited us was that we now had a way to visualise how two already known proteins interact together with this new factor to form a trimer.”
The scientists validated the AI predictions through experiments in living organisms– from the model organism zebrafish to mice, as well as human cells. They confirmed not only that this trimer forms in cells, but also that it exists across different species from different vertebrate groups—and when it fails to form, it makes the animal sterile. “This trimer is anchored at the sperm’s cell membrane, and two of the three proteins form the binding site for the egg protein in zebrafish,” continues Blaha.
The complex on the sperm’s membrane was found to interact with the zebrafish egg's gatekeeper Bouncer, located on the egg’s surface. Bouncer serves as a lock that only grants access to the egg with the right key, just like its mammalian counterpart Juno.
The sperm trimeric complex is evolutionarily conserved across vertebrates, while the interacting egg proteins have changed in different species to mediate the binding of sperm and egg.
“The identification of this complex of three proteins is a major step forward,” says Andrea Pauli. “The fact that it was maintained over millions of years of evolution shows just how important this lock-and-key process is. But what is really surprising is that the conserved sperm trimer uses evolutionarily unrelated egg proteins to dock onto the surface of the egg—evolutionary diversity resulting in a universal mechanism, right at the beginning of life!”
Original Publication
Victoria E. Deneke*#, Andreas Blaha*, Yonggang Lu, Johannes P. Suwita, Jonne M. Draper, Clara S. Phan, Karin Panser, Alexander Schleiffer, Laurine Jacob, Theresa Humer, Karel Stejskal, Gabriela Krssakova, Elisabeth Roitinger, Dominik Handler, Maki Kamoshita, Tyler D.R. Vance, Xinyin Wang, Joachim M. Surm, Yehu Moran, Jeffrey E. Lee, Masahito Ikawa, Andrea Pauli#. "A conserved fertilization complex bridges sperm and egg in vertebrates." Cell, DOI: 10.1016/j.cell.2024.09.035
*These authors contributed equally. #Corresponding authors.
About the Vienna BioCenter PhD Program
Much of the work underlying this publication was done by a doctoral student of the Vienna BioCenter PhD Program. Are you interested in a world-class career in molecular biology? Find out more: https://training.vbc.ac.at/phd-program/
About the IMP at the Vienna BioCenter
The Research Institute of Molecular Pathology (IMP) in Vienna is a basic life science research institute largely sponsored by Boehringer Ingelheim. With over 220 scientists from 40 countries, the IMP is committed to scientific discovery of fundamental molecular and cellular mechanisms underlying complex biological phenomena. The IMP is part of the Vienna BioCenter, one of Europe’s most dynamic life science hubs with 2,800 people from over 80 countries in six research institutions, two universities, and 35 biotech companies. www.imp.ac.at, www.viennabiocenter.org
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