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The transcription factor Otx2 acts as a negative switch in the regulation of transition from naive to primed pluripotency in mouse pluripotent stem cells. However, the molecular features and function of porcine OTX2 have not been well elucidated in porcine-induced pluripotent stem cells (piPSCs). By studying high-throughput transcriptome sequencing and interfering endogenous  OTX2  expression, we demonstrate that OTX2 is able to downgrade the self-renewal of piPSCs.  OTX2  is highly expressed in porcine brain, reproductive tissues, and preimplantation embryos, but is undetectable in fibroblasts and most somatic tissues. However, the known piPSC lines reported previously produced different levels of OTX2 depending on the induction procedures and culture conditions. Overexpression of porcine OTX2 can reduce the percentage of alkaline phosphatase-positive colonies and downregulate  NANOG  and  OCT4  expression. In contrast, knockdown of  OTX2  can significantly increase endogenous expressions of  NANOG ,  OCT4 , and  ESRRB , and stabilize the pluripotent state of piPSCs. On the other hand, NANOG can directly bind to the  OTX2  promoter as shown in ChIP-seq data and repress  OTX2  promoter activity in a dose-dependent manner. These observations indicate that OTX2 and NANOG can form a negative feedback circuitry to regulate the pluripotency of porcine iPS cells.

OTX2 impedes self–renewal of porcine iPS cells through downregulation of NANOG expression