The egg must be pre-equipped with the cellular machinery necessary to facilitate successful fertilization and, in oviparous species, to sustain and protect the embryo throughout development. An integral component of this early life support system is the thousands of maternally derived cortical granules (CG) which initiate the expansion of the perivitelline space upon egg activation/fertilization, facilitate prevention of polyspermy, and provide the embryo with the necessary componentry to develop a system of innate immunity. While the importance of CGs for successful fertilization is often recited, little is known regarding their composition. For this reason, we sought to identify candidate CG proteins from the perivitelline space of zebrafish eggs.
Ovulated eggs were collected from zebrafish and subjected to activation by exposure to water. Perivitelline fluid was aspirated and collected as a pooled sample until approximately 50 μl was acquired (~150 eggs). The pooled sample was processed for LC-MS-based protein profiling and the resulting peptide sequences were subjected to database-dependent protein identification, utilising UniProt’s zebrafish protein database.
Our preliminary analysis detected 78 proteins, each represented by at least two unique peptides, in the perivitelline fluid of zebrafish eggs, of which 41 proteins were identified as candidate CG proteins. Seven proteins have been selected for further interrogation by immunohistochemistry, quantitative PCR, and/or antibody-mediated knock-down to further characterize zebrafish CG and shed further light on their functional significance during fertilization and early embryogenesis.