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

Abstract

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.

Cell Type-Specific Expression Profile and Signaling Requirements in Early Hematopoietic Reprogramming

“…Interestingly, c-Myc, which is a key gene known to significantly impact TGF-b signaling, was expressed at a higher level in hematopoietic cells than in fibroblasts, as shown by our own RNA-seq data ( Supplementary Fig. S9) and an independent public dataset (GSE36290) [28]. This result is supportive of earlier findings in which a TGF-b inhibitor much more significantly promoted reprogramming in the absence than in the presence of c-Myc [25,26].…”

Unique features of mutations revealed by sequentially reprogrammed induced pluripotent stem cells

“…2). Moreover, in a recent study, we have demonstrated that the core histone modifications (H3K4me2, H3K4me3, H3K27me3) are comparable among iPSC lines that can produce all-iPSC mice 24 . Taken together, we conclude that although alliPSC mice can be produced up to six generations, their viability decreases with increasing generations, and epigenetic effects might not be a major cause of this reduction in viability.…”

“…MeDIP-seq and RNA-seq of all eight sequential iPSC lines were conducted as previously described 24 . In brief, the genomic DNA was sonicated to 100-500 bp and adaptor was then ligated to the end of the DNA fragments according to the paired-end DNA sample prep kit (Illumina).…”

microTSS: accurate microRNA transcription start site identification reveals a significant number of divergent pri-miRNAs

“…ESC small-RNA-Seq data were derived from the study of Chang et al 36 (GSM886478). Following pre-processing, adapter-trimmed reads were aligned against known human mature miRNA sequences (miRBase v20 (ref.…”