The Mechanism Of Maintaining Pluripotency In Pig Induced Stem Cells

The wide application of pig disease model has caused a surge of interest in the study ofderivation of pig induced pluripotent cells (iPSCs). While major progress has been made indefining the molecular networks that control pluripotentcy in human and mouse, themechanism underlying the maintainence of pluripotency in pig remain largely unknown.1. Comparative Gene Expression Signature of Pig, Human and Mouse InducedPluripotent Stem Cell Lines Reveals Insight into Pig Pluripotency Gene NetworksPig induced pluripotent stem cells (piPSCs) have significant biomedical and agriculturalapplications. In this study, we assessed the transcriptome profile of multiple piPSC linesderived from different laboratories worldwide and compared them to pig somatic, embryonicstem cells and mouse and human iPSCs to determine the molecular signaling pathways thatmight play a central role in authentic piPSCs. The results demonstrated that the up-regulationof endogenous epithelial cells adhesion molecule (EpCAM) was correlated with thepluripotent state of pig pluripotent cells, which could be utilized as a marker for evaluatingpig cell reprogramming. Comparison of key signaling pathways including JAK-STAT,NOTCH, TGFB1, WNT and VEGF in pig, human and mouse iPSCs showed that the coretranscriptional network to maintain pluripotency and self-renewal in pig were different fromthat in mouse, but had significant similarities to human. Pig iPSCs, which lacked expressionof specific na ve state markers KLF2/4/5and TBX3, but expressed the primed state markers ofOtx2and Fabp7, share defining features with human ESCs and mouse EpiSCs. The cluster ofimprinted genes delineated by the delta-like homolog1gene and the type III iodothyroninedeiodinase gene (DLK1-DIO3) were silenced in piPSCs as previously seen in mouse iPSCswhich failed to support the development of entirely iPSC-derived animals. These keydifferences in na ve state gene and imprinting gene expression suggests that in general piPSCsmay be more similar to primed state cells. The primed state of these cells may potentiallyexplain the rare ability of piPSCS to generate chimeras and cloned offspring.2. Comprehensive transcriptome analysis of pig induced pluripotent stem cells byhigh-throughput sequencingWe performed genome-wide analysis of gene expression profiling by RNA seq. Weidentified mRNA and miroRNA transcripts that were specifically expressed in pig iPSCs. Our analysis identify the genes upregulated in pig iPS compared with somatic cells and also thedifferentially expressed genes between pig iPSCs under different culture medium. Thesegenes may led to the identification of key factors which maintain or confer the pluripotentnature of pig iPSCs. We then pursued comprehensive bioinformatics analyses, includingfunctional annotation of the generated data within the context of biological pathways, touncover novel biological functions associated with maintenance of pluripotency in pig. Thisresult supports that pig iPS have transcript profiles linked to “ribosome”,“chromatinremodeling”, and genes involved in “cell cycle “that may be critical to maintain theirpluripotency, plasticity, and stem cell function. Our analysis demonstrate the key role of RNAsplicing in regulating the pluripotency phenotype of pig cells. Specifically, the data indicatedistinctive expression pattern for SALL4spliced variants in different pig cell types andhighlight the necessity of defining the type of SALL4when addressing the expression of thisgene in pig cells. At last, we profile the dynamic gene expression signature of pluripotentgenes in the preimplantation development embyo of pig. The resulting comprehensive dataallowed us to compare various different subsets of pig pluripotent cells, and we identifiedseveral genes and miRNAs that had not previously been reported to characterize pig iPS cells.This information provided by our anlaysis will ultimately advance the efforts at generatingstable na ve pluripotency in pig cells.