Proteomic analysis of porcine oocytes during in vitro maturation reveals essential role for the ubiquitin C-terminal hydrolase-L1

• DOI: 10.1530/rep-07-0079
• Publication Date: 2007-09-21T14:18:44Z
• Authors: Andrej Susor; Zdenka Ellederova; Lucie Jelinkova; Petr Halada; Daniel Kavan; Michal Kubelka; Hana Kovarova
• Keywords: 0301 basic medicine; Proteasome Endopeptidase Complex; Indoles; Swine; Ubiquitin; Spectrum Analysis; Egg Proteins; Immunoblotting; Myelin Basic Protein; Fertilization in Vitro; Meiosis; 03 medical and health sciences; CDC2 Protein Kinase; Oximes; Oocytes; Animals; Electrophoresis, Gel, Two-Dimensional; Female; Protein Kinases; Ubiquitin Thiolesterase

In this study, we performed proteomic analysis of porcine oocytes during in vitro maturation. Comparison of oocytes at the initial and final stages of meiotic division characterized candidate proteins that were differentially synthesized during in vitro maturation. While the biosynthesis of many of these proteins was significantly decreased, we found four proteins with increased biosynthetic rate, which are supposed to play an essential role in meiosis. Among them, the ubiquitin C-terminal hydrolase-L1 (UCH-L1) was identified by mass spectrometry. To study the regulatory role of UCH-L1 in the process of meiosis in pig model, we used a specific inhibitor of this enzyme, marked C30, belonging to the class of isatin O-acyl oximes. When germinal vesicle (GV) stage cumulus-enclosed oocytes were treated with C30, GV breakdown was inhibited after 28 h of culture, and most of the oocytes were arrested at the first meiosis after 44 h. The block of metaphase I–anaphase transition was not completely reversible. In addition, the inhibition of UCH-L1 resulted in elevated histone H1 kinase activity, corresponding to cyclin–dependent kinase(CDK1)–cyclin B1 complex, and a low level of monoubiquitin. These results supported the hypothesis that UCH-L1 might play a role in metaphase I–anaphase transition by regulating ubiquitin-dependent proteasome mechanisms. In summary, a proteomic approach coupled with protein verification study revealed an essential role of UCH-L1 in the completion of the first meiosis and its transition to anaphase.