Reproduction Abstracts

Introduction: The female reproductive tract provides the optimal environment for gamete interaction and embryo development. Understanding the oviductal environment is important for reproducing the in vivo conditions in vitro. OVGP1 is the major non-serum oviductal protein with a variable protein activity depending on its regions. To investigate the potential role of the OVGP1 regions on its activity, full-length (prOVGP1, 527aa) and C-terminal truncated (prOVGP1TR, 481aa) recombinant histidine-tagged porcine OVGP1 proteins were expressed in mammalian cells.

Material and methods: Secreted proteins were purified by immobilized-metal affinity chromatography followed by size exclusion chromatography. Media from transfected cells were incubated with in vitro-matured (IVM) porcine oocytes. After 1 h, oocytes were used to measure the resistance of zona pellucida (ZP) to pronase digestion and imaged by confocal microscopy using anti-His antibody. In a second experiment, purified prOVGP1TR protein was incubated with IVM oocytes for 1 h. Oocytes were washed and transferred to fresh IVF medium for insemination.

Results and discussion: Recombinant proteins were detected by fluorescent signal bound to ZP. The ZP resistance to proteolytic digestion increased in the oocytes incubated with prOVGP1TR protein compared with prOVGP1. The oocytes exposed to prOVGP1TR showed a significant increase in the percentage of monospermy and a decrease in the percentage of penetration compared with control oocytes. These results reveal that preincubation of oocytes with truncated OVGP1 protein not only increases ZP resistance to proteolysis but also affects sperm-penetration ability suggesting that C-terminal region of OVGP1 modifies protein function. Supported by MINECO and FEDER (AGL2012-40180-C03-01-02).