High-throughput sequencing reveals the disruption of methylation of imprinted gene in induced pluripotent stem cells

• DOI: 10.1038/cr.2013.173
• Publication Date: 2013-12-31T15:55:02Z
• Authors: Gang Chang; Shuai Gao; Xinfeng Hou; Zijian Xu; Yanfeng Liu; Lan Kang; Yu Tao; Wenqiang Liu; Bo Huang; Xiaochen Kou; Jiayu Chen; Lei An; Kai Miao; Keqian Di; Zhilong Wang; Kun Tan; Tao Cheng; Tao Cai; Shaorong Gao; Jianhui Tian
• Keywords: 0301 basic medicine; Sequence Analysis, RNA; Induced Pluripotent Stem Cells; High-Throughput Nucleotide Sequencing; Nuclear Proteins; DNA Methylation; Cellular Reprogramming; Chromatin; Epigenesis, Genetic; Histones; Tetraploidy; Genomic Imprinting; Mice; 03 medical and health sciences; Animals; Female; RNA, Messenger; Transcriptome; Cells, Cultured; Embryonic Stem Cells; Gene Library

It remains controversial whether the abnormal epigenetic modifications accumulated in the induced pluripotent stem cells (iPSCs) can ultimately affect iPSC pluripotency. To probe this question, iPSC lines with the same genetic background and proviral integration sites were established, and the pluripotency state of each iPSC line was characterized using tetraploid (4N) complementation assay. Subsequently, gene expression and global epigenetic modifications of "4N-ON" and the corresponding "4N-OFF" iPSC lines were compared through deep sequencing analyses of mRNA expression, small RNA profile, histone modifications (H3K27me3, H3K4me3, and H3K4me2), and DNA methylation. We found that methylation of an imprinted gene, Zrsr1, was consistently disrupted in the iPSC lines with reduced pluripotency. Furthermore, the disrupted methylation could not be rescued by improving culture conditions or subcloning of iPSCs. Moreover, the relationship between hypomethylation of Zrsr1 and pluripotency state of iPSCs was further validated in independent iPSC lines derived from other reprogramming systems.