
Cellular reprogramming of somatic cells towards induced pluripotency is a multistep stochastic process mediated by the transcription factors Oct4, Sox2, Klf4 and c-Myc (OSKM), which orchestrate global epigenetic and transcriptional changes. We performed a large-scale analysis of integrated ChIP-seq, ATAC-seq and RNA-seq data and revealed the spatiotemporal highly dynamic pattern of OSKM DNA binding during reprogramming. We found that OSKM show distinct temporal patterns of binding to different classes of pluripotency-related enhancers. Genes involved in reprogramming are regulated by the coordinated activity of multiple enhancers, which are sequentially bound by OSKM for strict transcriptional control. Based on these findings, we developed an unbiased approach to identify Reprogramming Inducible Enhancers (RIEs), constructed enhancer-traps and isolated cells undergoing reprogramming in real-time. We used a representative RIE taken from the Upp1 gene fused to GFP and isolated cells at different time points during reprogramming and found that they have unique developmental capacities as they are reprogrammed with high efficiency due to their distinct molecular signatures. In conclusion, our experiments have led to the development of an unbiased method to identify and isolate reprogrammable cells in real-time by exploiting the functional dynamics of OSKM, which can be used as efficient reprogramming biomarkers.
SOXB1 Transcription Factors, Induced Pluripotent Stem Cells, Kruppel-Like Transcription Factors, Cellular Reprogramming, Article, Proto-Oncogene Proteins c-myc, Kruppel-Like Factor 4, Mice, Enhancer Elements, Genetic, Cellular Reprogramming/genetics; Kruppel-Like Factor 4/metabolism; Animals; Enhancer Elements, Genetic; Mice; Octamer Transcription Factor-3/genetics; Octamer Transcription Factor-3/metabolism; Induced Pluripotent Stem Cells/metabolism; Induced Pluripotent Stem Cells/cytology; SOXB1 Transcription Factors/genetics; SOXB1 Transcription Factors/metabolism; Kruppel-Like Transcription Factors/genetics; Kruppel-Like Transcription Factors/metabolism; Transcription Factors/metabolism; Transcription Factors/genetics; Proto-Oncogene Proteins c-myc/metabolism; Proto-Oncogene Proteins c-myc/genetics; Chromatin Immunoprecipitation Sequencing/methods; OSKM; cellular reprogramming; chromatin structure; enhancers; genomics; iPSCs; transcription factors; transcriptional regulation, Animals, Chromatin Immunoprecipitation Sequencing, Octamer Transcription Factor-3, Transcription Factors
SOXB1 Transcription Factors, Induced Pluripotent Stem Cells, Kruppel-Like Transcription Factors, Cellular Reprogramming, Article, Proto-Oncogene Proteins c-myc, Kruppel-Like Factor 4, Mice, Enhancer Elements, Genetic, Cellular Reprogramming/genetics; Kruppel-Like Factor 4/metabolism; Animals; Enhancer Elements, Genetic; Mice; Octamer Transcription Factor-3/genetics; Octamer Transcription Factor-3/metabolism; Induced Pluripotent Stem Cells/metabolism; Induced Pluripotent Stem Cells/cytology; SOXB1 Transcription Factors/genetics; SOXB1 Transcription Factors/metabolism; Kruppel-Like Transcription Factors/genetics; Kruppel-Like Transcription Factors/metabolism; Transcription Factors/metabolism; Transcription Factors/genetics; Proto-Oncogene Proteins c-myc/metabolism; Proto-Oncogene Proteins c-myc/genetics; Chromatin Immunoprecipitation Sequencing/methods; OSKM; cellular reprogramming; chromatin structure; enhancers; genomics; iPSCs; transcription factors; transcriptional regulation, Animals, Chromatin Immunoprecipitation Sequencing, Octamer Transcription Factor-3, Transcription Factors
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