According to the World Health Organization (WHO) around 15% of couples worldwide face infertility, with some cases linked to mutations in genes responsible for the egg coat, or zona pellucida (ZP). The ZP is a crucial extracellular matrix for egg growth, fertilization, and embryo protection until implantation in the uterus. It comprises proteins that form a thick filamentous ‘mesh’ around the egg. Mutations in ZP proteins can lead to defects or absence of the mesh. Although the prevalence of pathogenic mutations in ZP genes among infertile women remains uncertain, due to the lack of comprehensive genome sequencing, several types of such mutations have been documented. After fertilization, the egg releases a proteolytic enzyme that cleaves the N-terminal region of ZP2, a glycoprotein and major subunit of ZP filaments. This makes the egg coat ‘harder’ and was thought to inactivate sperm binding, but a molecular mechanism remained elusive.
In this study we investigated the structure of ZP2 before and after cleavage, and analysed how this modifies the overall ZP mesh structure [1]. With this goal, different ZP2 and egg filaments structures were analysed using X-ray crystallography and cryo-electron microscopy. These revealed that cleavage of the part of ZP2 that protrudes from the egg filament promotes the establishment of new interactions with the same part of other ZP2 molecules. These new interactions induce an extensive network of cross-links that moves the filaments closer to each other, stiffening the egg coat by tightening its mesh. In this manner polyspermy is not avoided by stopping other sperm to bind to the ZP, but rather by hindering sperm penetration (Figure). The findings have important implications for reproductive medicine, offering insights into human ZP gene mutations linked to female infertility and potential applications in non-hormonal contraceptive development.
The ZP2 and egg coat filament structures were determined by combining a multidisciplinary approach that used X-ray crystallography at the ESRF, Diamond Light Source and BESSY II, as well as cryo-electron microscopy at SciLifeLab in Stockholm. Prediction from AlphaFold2 were used to model the human ZP filaments.
Daniele de Sanctis (ESRF) and Luca Jovine (Karolinska Institutet)
[1] Nishio S, Emori C, Wiseman B, Fahrenkamp D, Dioguardi E et al. (2024) Cell, 187, 1440-1459
Figure: Graphical representation on egg coating stiffening after fertilization and ZP2 cleavage (adapted from [1])
