Department of Animal Genetics, Breeding and Reproduction, College of Animal Science and Technology, Nanjing Agricultural University, Nanjing 210095, China.
Anhui Province Key Laboratory of Local Livestock and Poultry, Genetical Resource Conservation and Breeding, College of Animal Science and Technology, Anhui Agricultural University, Hefei 230036, China.
Int J Mol Sci. 2022 Oct 8;23(19):11941. doi: 10.3390/ijms231911941.
Porcine-induced pluripotent stem cells (piPSCs) are of great significance to animal breeding and human medicine; however, an important problem is that the maintenance of piPSCs mainly depends on exogenous expression of pluripotent transcription factors (TFs), and germline transmission-competent piPSCs have not yet been successfully established. In this study, we explore the defect of epigenetic reprogramming during piPSCs formation, including chromatin accessibility, DNA methylation, and imprinted gene expression, with high-throughput sequencing (ATAC-seq, WGBS, RNA-seq, and Re-seq) methods. We found the somatic features were successfully silenced by connecting closed chromatin loci with downregulated genes, while DNA methylation has limited effects on somatic silence. However, the incomplete chromatin remodeling and DNA demethylation in pluripotency genes hinder pluripotent activation, resulting in the low expression of endogenous pluripotency genes. In addition, the expression of potential imprinted genes was abnormal, and many allelic-biased expressed genes in porcine embryonic fibroblasts (PEFs) were erased, accompanied by establishment of new allelic-biased expressed genes in piPSCs. This study reveals the aberrant epigenetic reprogramming during dox-dependent piPSCs formation, which lays the foundation for research of porcine-iPSC reprogramming and genome imprinting.
猪诱导多能干细胞（piPSCs）对动物育种和人类医学具有重要意义；然而，一个重要的问题是 piPSCs 的维持主要依赖于多能转录因子（TFs）的外源表达，并且还没有成功建立具有种系传递能力的 piPSCs。在这项研究中，我们使用高通量测序（ATAC-seq、WGBS、RNA-seq 和 Re-seq）方法探索了 piPSCs 形成过程中表观遗传重编程的缺陷，包括染色质可及性、DNA 甲基化和印迹基因表达。我们发现，通过将封闭染色质位点与下调基因连接，可以成功沉默体细胞特征，而 DNA 甲基化对体细胞沉默的影响有限。然而，在多能性基因中，不完全的染色质重塑和 DNA 去甲基化阻碍了多能性的激活，导致内源性多能性基因的低表达。此外，潜在印迹基因的表达异常，猪胚胎成纤维细胞（PEFs）中许多等位基因偏性表达的基因被抹去，同时在 piPSCs 中建立了新的等位基因偏性表达的基因。这项研究揭示了 dox 依赖性 piPSCs 形成过程中异常的表观遗传重编程，为猪-iPSC 重编程和基因组印迹的研究奠定了基础。
