Molecular characterization and polyadenylation-regulated expression of cyclin B1 and Cdc2 in porcine oocytes and early parthenotes.
Meiotic maturation of mammalian oocytes is controlled by the maturation/M-phase promotion factor (MPF), a complex of Cdc2 kinase and cyclin B protein. To better understand the molecular mechanism of oocyte maturation, we characterized porcine cyclin B1 and Cdc2 genes, both of which are widely expressed in pig tissues. We further analyzed their expression profiles during in vitro maturation of pig oocyte and early embryonic development at both the mRNA and protein level. Two isoforms of cyclin B1, comprising the same open reading frame but differing in 3'-UTR length, were identified. Cyclin B1 transcripts was up-regulated after 30 hr of maturation, while Cdc2 mRNA levels were unchanged during maturation except for a sharp decline at 44 hr. Cyclin B1 protein synthesis increased with oocyte maturation. Cdc2 protein expression was relatively low during 0-18 hr, followed by a higher level of expression up to 44 hr of maturation. Poly(A)-test PCR clearly revealed that both cyclin B1 isoforms underwent cytoplasmic polyadenylation starting around 18-24 hr during maturation, while a substantial de-adenylation and degradation of Cdc2 isoforms were observed in metaphase II oocytes and during embryo development after parthenogenetic activation. Porcine MII oocytes derived from small follicles (< or = 3 mm) and bad quality 2-cell parthenotes showed lower developmental competence and lower levels of cyclin B1 protein, and Cdc2 mRNA or both gene mRNAs, respectively, compared to their control counterparts. These results suggested that cyclin B1 was regulated posttranscriptionally by cytoplasmic polyadenylation during porcine oocyte maturation. Further, the decreased expression of maternal cyclin B1 and Cdc2 at the mRNA or protein level in developmentally incompetent oocytes and embryos was responsible for, at least in part, a profound defect in further embryonic development.
Zhang
Ding-Xiao
DX
Department of Animal Sciences, Chungbuk National University, Chungbuk, South Korea.
Cui
Xiang-Shun
XS
Kim
Nam-Hyung
NH
D020413:3' Untranslated Regions; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D016203:CDC2 Protein Kinase / Q000235:genetics / Q000378:metabolism*; D056744:Cyclin B1 / Q000235:genetics / Q000378:metabolism*; D030541:Databases, Genetic; D004622:Embryo, Mammalian / Q000502:physiology; D020224:Expressed Sequence Tags; D005260:Female; D018507:Gene Expression Regulation, Developmental; D006801:Humans; D008969:Molecular Sequence Data; D009865:Oocytes / Q000166:cytology / Q000502:physiology*; D010312:Parthenogenesis / Q000502:physiology*; D026723:Polyadenylation; D011247:Pregnancy; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism*; D016415:Sequence Alignment; D013552:Swine; D014018:Tissue Distribution
D020413:3' Untranslated Regions; D056744:Cyclin B1; D020033:Protein Isoforms; D016203:CDC2 Protein Kinase