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Cell Rep. 2019 May 7;27(6):1726-1741.e5. doi: 10.1016/j.celrep.2019.04.056.

Defining Reprogramming Checkpoints from Single-Cell Analyses of Induced Pluripotency.

Author information

1
Wisconsin Institute for Discovery, University of Wisconsin-Madison, Madison, WI, USA.
2
Department of Human Oncology, University of Wisconsin-Madison, Madison, WI, USA.
3
Wisconsin Institute for Discovery, University of Wisconsin-Madison, Madison, WI, USA; Department of Biomedical Informatics, University of Wisconsin-Madison, Madison, WI, USA.
4
Wisconsin Institute for Discovery, University of Wisconsin-Madison, Madison, WI, USA; Department of Cell and Regenerative Biology, University of Wisconsin-Madison, Madison, WI, USA. Electronic address: rsridharan2@wisc.edu.

Abstract

Elucidating the mechanism of reprogramming is confounded by heterogeneity due to the low efficiency and differential kinetics of obtaining induced pluripotent stem cells (iPSCs) from somatic cells. Therefore, we increased the efficiency with a combination of epigenomic modifiers and signaling molecules and profiled the transcriptomes of individual reprogramming cells. Contrary to the established temporal order, somatic gene inactivation and upregulation of cell cycle, epithelial, and early pluripotency genes can be triggered independently such that any combination of these events can occur in single cells. Sustained co-expression of Epcam, Nanog, and Sox2 with other genes is required to progress toward iPSCs. Ehf, Phlda2, and translation initiation factor Eif4a1 play functional roles in robust iPSC generation. Using regulatory network analysis, we identify a critical role for signaling inhibition by 2i in repressing somatic expression and synergy between the epigenomic modifiers ascorbic acid and a Dot1L inhibitor for pluripotency gene activation.

PMID:
31067459
DOI:
10.1016/j.celrep.2019.04.056
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