Analyses for the regulation mechanisms of protein synthesis and degradation which is aimed for the improvement of the availability in live stock oocyte for the developmental technology

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I analyzed the molecular mechanisms for regulating the protein synthesis and degradation during normal development in mammalian oocytes/early embryos. Especially, using maturing porcine oocytes as examples, I focused on Aurora A as the regulator of protein synthesis and on cdc20 and cdh1, which were the activator of anaphase promoting complexes (APC: a ubiquitine ligase working in a ubiquitine/proteasome system), as the regulator of protein degradation. During 2005, I succeeded for the cloning of these genes in the pig and registered them in a gene bank.The Aurora A protein level in porcine oocytes was about 100 times higher than that in somatic cells, indicating the importance of its function in the oocyte maturation. The injection of the mRNA for a constitutive active mutant of Aurora A into porcine immature oocytes accelerated the synthesis of cyclin B and subsequent meiotic resumption. Conversely, the injection of the antisense RNA of Aurora A significantly inhibited the meiotic re … More sumption under the condition of Aurora A depletion in porcine oocytes. These are the first results showing the importance of Aurora A on the protein synthesis during porcine oocyte meiosis.I also injected the antisense RNAs of cdc20 and cdh1 into porcine immature oocytes. These experiments revealed that the inhibition of cdc20 expression induced the arrest at the first meiotic metaphase in porcine oocytes. In these oocytes, the degradation of cyclin B was prevented and MPF activity was maintained at a high level. In contrast, the inhibition of cdh1 expression increased the cyclin B accumulation at the GV stage and accelerated the meiotic resumption, indicating the cdh1-dependent cyclin B degradation during meiotic arrest in porcine oocytes. These results suggest the important functions of cdh1 and cdc20 through cyclin B degradation, the inhibition of premature meiotic resumption and the first meiosis/the second meiosis transition, respectively. Furthermore, I found that the cdh1 overexpression inhibited the meiotic resumption and cdc20 might be a substrate of cdh1/APC. The present study extended our understandings in the regulation of protein synthesis and degradation in mammalian oocytes. Less