Staged miRNA re-regulation patterns during reprogramming

Background: MiRNAs often operate in feedback loops with transcription factors and represent a key mechanism for fine-tuning gene expression. In transcription factor-induced reprogramming, miRNAs play a critical role, however, detailed analyses of miRNA expression changes during reprogramming at the level of deep sequencing have not been previously reported. Results: We use four factor reprogramming to induce pluripotent stem cells from mouse fibroblasts and isolate FACS-sorted Thy1-and SSEA1+ intermediates and Oct4-GFP+ induced pluripotent stem cells (iPSCs). Small RNAs from these cells, and two partial-iPSC lines, another iPSC line, and mouse embryonic stem cells (mES cells) were deep sequenced. A comprehensive resetting of the miRNA profile occurs during reprogramming; however, analysis of miRNA co-expression patterns yields only a few patterns of change. Dlk1-Dio3 region miRNAs dominate the large pool of miRNAs experiencing small but significant fold changes early in reprogramming. Overexpression of Dlk1-Dio3 miRNAs early in reprogramming reduces reprogramming efficiency, suggesting the observed downregulation of these miRNAs may contribute to reprogramming. As reprogramming progresses, fewer miRNAs show changes in expression, but those changes are generally of greater magnitude. Conclusions: The broad resetting of the miRNA profile during reprogramming that we observe is due to small changes in gene expression in many miRNAs early in the process, and large changes in only a few miRNAs late in reprogramming. This corresponds with a previously observed transition from a stochastic to a more deterministic signal.