Bonnet Monkey (Macaca radiata) Ovaries, Like Human Oocytes, Express Four Zona Pellucida Glycoproteins

Abstract

Recent studies document that the zona pellucida matrix of human oocytes is composed of four glycoproteins designated as ZP1, ZP2, ZP3, and ZP4 instead of three as proposed in mouse model. In the present study, investigations were carried out to find the presence of the fourth ZP glycoprotein in the ovaries of non-human primates namely bonnet monkey (Macaca radiata). Employing total RNA isolated from bonnet monkey ovaries, the complementary deoxyribonucleic acid (cDNA) encoding bonnet monkey ZP1 (up to furin cleavage site) was successfully amplified by reverse transcribed polymerase chain reaction (RT-PCR). The deduced amino acid (aa) sequence of bonnet monkey ZP1 revealed 96.0% identity with human ZP1. The 21 cysteine residues present in bonnet monkey ZP1 were conserved in human, mouse, rat, quail, and chicken. Simultaneously, polyclonal antibodies were generated in mice against synthetic peptides corresponding to human ZP1 (P1, 137-150 aa; P2, 223-235 aa; P3, 237-251 aa; P4, 413-432 aa; and P5, 433-451 aa). Employing anti-peptide antibodies that were devoid of cross-reactivity as determined by ELISA with human/bonnet monkey recombinant ZP2, ZP3, and ZP4, the presence of ZP1 in the ovaries of bonnet monkey and human oocytes was demonstrated by indirect immunofluorescence. The antibodies against peptides P3 and P4 reacted only with the ZP of bonnet monkey ovaries and not other ovarian associated cells. The data presented in this manuscript provide evidence, for the first time, that the bonnet monkey ZP matrix is composed of four glycoproteins.