Distinct MicroRNA Expression Signatures of Porcine Induced Pluripotent Stem Cells under Mouse and Human ESC Culture Conditions

It is well known that microRNAs play a very important role in regulating reprogramming, pluripotency and cell fate decisions. Porcine induced pluripotent stem cells (piPSCs) are now available for studying the pluripotent regulation network in pigs. Two types of piPSCs have been derived from human and mouse embryonic stem cell (ESC) culture conditions: hpiPSCs and mpiPSCs, respectively. The hpiPSCs were morphologically similar to human ESCs, and the mpiPSCs resembled mouse ESCs. However, our current understanding of the role of microRNAs in the development of piPSCs is still very limited. Here, we performed small RNA sequencing to profile the miRNA expression in porcine fibroblasts (pEFs), hpiPSCs and mpiPSCs. There were 22 differential expressed (DE) miRNAs down-regulated in both types of piPSCs compared with pEFs, such as ssc-miR-145-5p and ssc-miR-98. There were 27 DE miRNAs up-regulated in both types of piPSCs compared with pEFs. Among these up-regulated DE miRNAs in piPSCs, ssc-miR-217, ssc-miR-216, ssc-miR-142-5p, ssc-miR-182, ssc-miR-183 and ssc-miR-96-5p have much higher expression levels in mpiPSCs, while ssc-miR-106a, ssc-miR-363, ssc-miR-146b, ssc-miR-195, ssc-miR-497, ssc-miR-935 and ssc-miR-20b highly expressed in hpiPSCs. Quantitative stem-loop RT-PCR was performed to confirm selected DE miRNAs expression levels. The results were consistent with small RNA sequencing. Different expression patterns were observed for key miRNA clusters, such as the miR-17-92 cluster, the let-7 family, the miR-106a-363 cluster and the miR-182-183 cluster, in the mpiPSCs and hpiPSCs. Novel miRNAs were also predicted in this study, including a putative porcine miR-302 cluster: ssc_38503, ssc_38503 and ssc_38501 (which resemble human miR-302a and miR-302b) found in both types of piPSCs. The miR-106a-363 cluster and putative miR-302 cluster increased the reprogramming efficiency of pEFs. The study revealed significant differences in the miRNA signatures of hpiPSCs and mpiPSCs under different pluripotent states that were derived from different culture conditions. These differentially expressed miRNAs may play important roles in pluripotent regulation in pigs, and this information will facilitate the understanding of the mechanism of pluripotency in pigs.

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