219 EXPRESSION PATTERN OF THE SUB-CORTICAL MATERNAL COMPLEX IN OVINE OOCYTES AND PRE-IMPLANTATION EMBRYOS

The sub-cortical maternal complex (SCMC) is a multi-protein complex located in the sub-cortex of the oocyte. In mouse, it assembles during oocyte growth and is essential for zygotes to progress beyond the first embryonic cell divisions (Li et al., 2008). At least 4 proteins contribute to the complex: oocyte expressed protein (OOEP), maternal antigen that embryo requires (MATER), transducin-like enhancer of split 6 (TLE6), and ES cell associated transcript 1 (ECAT1), all encoded by maternal effect genes. In mouse, the relative transcripts are degraded during meiotic maturation and ovulation, whereas the SCMC proteins persist in the early embryo. Whereas MATER expression has been studied in several species, the existence of the genes encoding the other components has been assessed in few mammalian species and their pattern of expression during pre-implantation development has been analysed only in mouse (Li et al. 2008 Dev. Cell 15, 416–425). In a previous work (Bebbere et al. 2008 Reprod. Fertil. Dev. 20, 908–915), we assessed MATER existence and pattern of expression in the ovine species. The aim of the present work was to assess the existence of OOEP, TLE6, and FILIA in the ovine species and to analyse the expression pattern of the 4 genes in the oocytes and during pre-implantation embryo development. Total RNA was isolated and reverse transcribed from pools of immature (GV) and in vitro matured (IVM) metaphase II (MII) oocytes, from in vitro fertilized and cultured (IVFC) embryos at the 2-, 4-, 8-, and 16-cell stage and from blastocysts. Three pools of 10 oocyte/embryos were analysed for each class. Primers were designed on the basis of the sequences conserved among orthologs and amplify intron-spanning regions. The PCR products were sequenced, and the alignment, performed with BLASTn, confirmed the homology with the orthologous genes present in public databases. Real-time PCR analysis revealed that all 4 transcripts are present at its highest level in the GV oocyte but decrease during embryo pre-implantation development with a gene-specific pattern. Conversely to the pattern of expression observed in mouse, all 4 transcripts persisted until the 8-cell stage embryo, disappearing only at the 16-cell stage. No transcripts were detected at the blastocyst stage. This study confirms the existence of transcripts related to SCMC also in the ovine species, but highlights species-specific patterns of expression in the 2 species, possibly related to the different time of activation of the embryo genome in mouse and in sheep. The observed expression patterns suggest an involvement of the protein complex in oocyte maturation and in the very first phases of life, possibly in the transition from the maternal to embryonic program of development.