Poly(A) RNA is reduced by half during bovine oocyte maturation but increases when meiotic arrest is maintained with CDK inhibitors

Abstract

Variations in the amount of different RNA species were investigated during in vitro maturation of bovine oocytes. Total RNA content was estimated to be 2 ng before meiosis, and after meiosis resumption, no decrease was observed. Ribosomal RNA did not appear to be degraded either, whereas poly(A) RNA was reduced by half after meiosis resumption, from 53 pg to 25 pg per oocyte. Real-time polymerase chain reaction was performed on growth and differentiation factor-9 (GDF-9), on cyclin B1, and on two genes implicated in the resistance to oxidative stress, glucose-6-phosphate-dehydrogenase (G6PD) and peroxiredoxin-6 (PRDX6). When these transcripts were reverse-transcribed with hexamers, the amplification results were not different before or after in vitro maturation. But when reverse transcription was performed with oligo(dT), amplification was dramatically reduced after maturation, except for cyclin B1 mRNA, implying deadenylation without degradation of three transcripts. Although calf oocytes have a lower developmental competence, their poly(A) RNA contents were not different from that of cow oocytes, nor were they differently affected during maturation. When bovine oocytes were maintained in vitro under meiotic arrest with CDK inhibitors, their poly(A) RNA amount increased, but this rise did not change the poly(A) RNA level once maturation was achieved. The increase could not be observed under transcription inhibition and, when impeding transcription and adenylation, the poly(A) RNA decreased to a level normally observed after maturation, in spite of the maintenance of meiotic arrest. These results demonstrate the importance of adenylation and deadenylation processes during in vitro maturation of bovine oocytes.