Time-resolved small-RNA sequencing identifies microRNAs critical for formation of embryonic stem cells from the inner cell mass of mouse embryos

Cells of the inner cell mass (ICM) acquire a unique ability for unlimited self-renewal during transition into embryonic stem cells (ESCs) in vitro, while preserving their natural multi-lineage differentiation potential. Several different pathways have been identified to play critical roles in the formation of ESCs but the function of non-coding RNAs in this process is poorly understood. Here, we describe several microRNAs (miRNAs) that are crucial for efficient generation of mouse ESCs from ICMs. Using small-RNA sequencing, we characterize dynamic changes in miRNA expression profiles during outgrowth of ICMs in a high-resolution, time-course dependent manner. We detected several waves of miRNA transcription during ESC formation, to which miRNAs from the imprinted Dlk1-Dio3 locus contribute extensively. Stringent in silico and functional analyses reveal that Dlk1-Dio3 locus-embedded miRNAs (miR-541-5p, miR-410-3p, and miR-381-3p), miR-183-5p and miR-302b-3p promote, while miR-212-5p and let-7d-3p inhibit efficient ICM transition to ESCs. Collectively, these findings offer new mechanistic insights into the role of miRNAs during ESC derivation.