Abstract
MicroRNAs (miRNAs) are small, noncoding RNAs that regulate gene expression after transcription. miRNAs are present in transcriptionally quiescent full-grown oocytes and preimplantation embryos that display a low level of transcription prior to embryonic genome activation. The role of miRNAs, if any, in preimplantation development is not known. The temporal pattern of expression of miRNAs during bovine preimplantation development was determined by small RNA-sequencing using eggs and preimplantation embryos (1-cell, 2-cell, 4-cell, 8-cell, 16-cell, morula, and blastocyst). Embryos cultured in the presence of α-amanitin, which permitted the distinguishing of maternal miRNAs from embryonic miRNAs, indicated that embryonic miRNA expression was first detected at the two-cell stage but dramatically increased during the morula and blastocyst stages. Targeting DGCR8 by a small-interfering RNA/morpholino approach revealed a role for miRNAs in the morula-to-blastocyst transition. Knockdown of DGCR8 not only inhibited expression of embryonically expressed miRNAs but also inhibited the morula-to-blastocyst transition. In addition, RNA-sequencing identified an increased relative abundance of messenger RNAs potentially targeted by embryonic miRNAs in DGCR8-knockdown embryos when compared with controls. Results from these experiments implicate an essential role for miRNAs in bovine preimplantation embryo development.