Selective degradation of cyclin B1 mRNA in rat oocytes by RNA interference (RNAi).

Abstract

Cyclic adenosine monophosphate (cAMP) keeps oocytes in meiotic arrest, thereby preventing activation of the key regulators of meiosis, p34cdc2/cyclin B1, (known as maturation-promoting factor (MPF)) and Erk 1 and 2, members of the mitogen-activated protein kinase (MAPK) family. The activity of MAPK in oocytes is upregulated by Mos. We previously demonstrated that Mos translation in rat oocytes is negatively regulated by a PKA-mediated cAMP action, which inhibits c-mos mRNA polyadenylation and is associated with the suppression of p34 cdc2 kinase. The goal of the present study was to provide definitive evidence that Mos translation is subjected to MPF regulation. In order to inhibit MPF activity, we employed the double-stranded (ds) RNA interference (RNAi) of gene expression. We demonstrated that the introduction of cyclin B1 dsRNA into rat oocytes selectively depleted the corresponding mRNA, further ablating its protein product. These oocytes, which exhibit low MPF activity, failed to elongate the c-mos mRNA poly(A) tail, did not accumulate Mos and were unable to activate MAPK. We conclude that an active MPF in rat oocytes is necessary for c-mos mRNA polyadenylation and Mos translation.