Proteins in the extraembryonic matrix of preimplantation rabbit embryos

Preimplantation embryos of several species are surrounded by an extraembryonic matrix (often simply named zona pellucida) until briefly before implantation. All signals of the early embryo-maternal dialogue have to pass this matrix and therefore are detectable inside. We investigated the protein pattern of the extraembryonic matrices of 3-6-day-old rabbit embryos by two-dimensional gel-electrophoresis. Using (35)S-methionine incorporation, embryonic proteins were labelled and could be distinguished from maternal proteins. Furthermore, the presence of three different proteins (insulin-like growth factor-binding protein-3, uteroglobin, haptoglobin) within the matrices of day-6 embryos was investigated by Western blot analysis. The pattern and numbers of protein spots detected was clearly dependent on the time of embryonic development. Of all proteins detected, 19.3% and 33% are of embryonic origin (day 5 and day 6, respectively). At day 4 the zona proteins are no longer detectable, reflecting the degradation of the zona pellucida. From day 4 to day 5 proteins detectable within the extraembryonic matrices increase enormously. This demonstrates that embryo-maternal signalling accelerates at least 2 days before implantation. Insulin-like growth factor-binding protein-3, uteroglobin and haptoglobin are part of the early signalling as shown by Western blot analysis. Insulin-like growth factor-binding protein-3 could be detected as one spot at 38 kDa pI 6.1, uteroglobin at 8 kDa pI 6.0 and haptoglobin as two spots/isoforms at 36/38 kDa pI 5.8 and pI 6.0. These results demonstrate that extraembryonic matrices serving as a mailbox are a valuable tool for investigating early embryo-maternal signalling.