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New Data Reveal Role of microRNAs in Induced Pluripotency

NEW YORK (GenomeWeb) – An international team of researchers has reported new data that reveals broad changes in microRNA expression during the reprogramming of somatic cells into induced pluripotent stem cells (iPSCs) and suggests that a subset of the small, non-coding RNAs support pluripotency while the cells transition between the two states.

Reprograming somatic cells into iPSCs involves the forced expression of four transcription factors — Oct4, Sox2, Klf4 and c-Myc, collectively known as OSKM — that reset their epigenetic state and yield characteristics similar to embryonic stem cells (ESCs).

Amid growing interest in harnessing iPSCs for drug discovery and regenerative medicine, a group of more than 50 researchers from labs in Canada, Australia, Korea, and Holland joined together to form the Project Grandiose Consortium to uncover the molecular basis and means that lead to pluripotency.

While exploring alternative outcomes of somatic reprogramming, the consortium identified an novel stable pluripotent state characterized by a class of "fuzzy" colony-forming cell lines, dubbed F-class, that can arise when OSKM expression is maintained at a high level, according to a report published last week in Nature Communications.

To examine the changes that occur in somatic cells as they move into this F-class state or a classic ESC-like state, the scientists used doxycycline-inducible OSKM expression in secondary murine embryonic fibroblasts and performed small RNA sequencing at different time points. They discovered dramatic changes in individual miRNA expression, particularly at the early and late stages of the cells' transition, but noted that the cells still retained miRNA pools of similar size relative to total RNA content, pointing to the importance of tight regulation of overall miRNA levels.

The consortium researchers also found that many pluripotency-related miRNAs were expressed as multiple isoforms, which "presents important implications for their biological function and reliable detection," they wrote. Still, there was little variation in expression of these isoforms between cell states, indicating that changes in miRNA processing bias are not characteristic of the reprogramming process.

When weighing their observations of miRNA expression shifts against the epigenome and RNA transcriptome data generated in parallel, the investigators found evidence of both transcriptional and post-transcriptional mechanisms driving changes in the small RNAs.

Further, miRNA transcription was commonly regulated by dynamic histone modification, while DNA methylation/demethylation consolidates these changes at multiple loci, according to the paper.

Notably, the researchers found that a deficiency of F-class cells in well-characterized pluripotency-associated miRNAs can be compensated for by higher expression of another functionally equivalent subset of miRNAs.

Overall, the report represents a comprehensive small RNA expression analysis of somatic cell reprogramming that reveals "complex patterns of regulated miRNA biogenesis to support distinct states of cellular pluripotency," the study's authors concluded.

Combined with the concordant long RNA transcriptome, global proteome, and CpG methylation data that have been generated by the Project Grandiose Consortium, the new work "provides a unique resource to study dynamic genome expression during iPSC generation," they added.