UC Davis

Little is known about the proteins involved in gamete plasma membrane interaction during mammalian fertilization, and even less is known about the molecules involved in livestock species, such as the pig. Affinity chromatography of porcine sperm and oocyte plasma membrane proteins followed by mass spectrometry to identify eluted proteins detected apolipoprotein E (APOE) and a 167 amino acid Jacalin-type lectin domain-containing protein (A0A287AVU8) as potential proteins involved in this interaction (Vazquez Morales, 2023). To further explore the potential involvement of porcine APOE and A0A287AVU8 in fertilization, this study utilized an antibody inhibition technique. APOE antibodies were incubated with oocytes and A0A287AVU8 antibody were incubated with sperm for 10 minutes each prior to gamete co-incubation with the final dilution of antibody during sperm-oocyte interaction being 5 ng/µl. Each of the three replicates contained approximately 50 oocytes per treatment. A reduced ratio of fertilization (P < 0.05) was observed following the inclusion of the first APOE antibody (amino acid peptide residues 101-200) compared to the control (no antibodies). A trend toward reduced fertilization (P = 0.10) was observed with a second APOE antibody (amino acid peptide residues 151-250). Isotype control rabbit IgG did not alter fertilization. No effect on fertilization ratio was observed following incubation with rabbit anti-A0A287AVU8 IgG (P > 0.05), compared to the control (rabbit IgG), or sodium azide-containing rabbit IgG. These results suggest that APOE on the oocyte surface may contribute to effective sperm-oocyte interaction during porcine fertilization. This might be a specific receptor-ligand interaction or a more general “sticky” interaction involving APOE. Alexa Fluor 594-conjugated secondary antibody successfully localized APOE on the plasma membrane of the oocyte, and A0A287AVU8 on the sperm’s acrosome and tail, intensively staining the extremity of the latter. These findings contribute to the overall understanding of porcine fertilization.