Abstract
MicroRNAs play a pivotal role in cellular maintenance, proliferation, and differentiation. They have also been implicated to play a key role in disease pathogenesis, and more recently, cellular reprogramming. Certain microRNA clusters can enhance or even directly induce reprogramming, while repressing key proteins involved in microRNA processing decreases reprogramming efficiency. Although microRNAs clearly play important roles in cellular reprogramming, it remains unknown whether microRNAs are absolutely necessary. We endeavored to answer this fundamental question by attempting to reprogram Dicer-null mouse embryonic fibroblasts (MEFs) that lack almost all functional microRNAs using a defined set of transcription factors. Transduction of reprogramming factors using either lentiviral or piggyBac transposon vector into two, independently derived lines of Dicer-null MEFs failed to produce cells resembling embryonic stem cells (ESCs). However, expression of human Dicer in the Dicer-null MEFs restored their reprogramming potential. Our study demonstrates for the first time that microRNAs are indispensable for dedifferentiation reprogramming.
Highlights
MicroRNAs have emerged as a new category of genes that influence many cellular processes including proliferation and differentiation. miRNAs are small, noncoding, single stranded RNAs usually 22 nucleotides long that can base pair with target mRNAs in the open reading frame or 39 untranslated region [1]. miRNAs downregulate target genes by inhibiting protein translation and destabilizing mRNAs via deadenylation [2]
DicerD/D Mouse Embryonic Fibroblasts Lacking miRNAs are Viable Despite Suppressed Proliferation To test whether miRNAs are necessary for reprogramming a somatic cell type into induced stem cell-like cells, we first generated mouse embryonic fibroblasts (MEFs) that lack almost all miRNAs
Reverse transcription-quantitative polymerase chain reaction revealed that the levels of most mature miRNAs tested were nearly 98% depleted by 6 dpi. (Fig 1D)
Summary
MicroRNAs (miRNAs) have emerged as a new category of genes that influence many cellular processes including proliferation and differentiation. miRNAs are small, noncoding, single stranded RNAs usually 22 nucleotides long that can base pair with target mRNAs in the open reading frame or 39 untranslated region [1]. miRNAs downregulate target genes by inhibiting protein translation and destabilizing mRNAs via deadenylation [2]. Despite these variations and optimizations, DicerD/D MEFs could not be reprogrammed into induced stem cell-like cells when either 4 TFs or 5 TFs were transduced 6 days after induction with Cre (Fig 2A), when all mature miRNAs were depleted.
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
