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Incomplete DNA methylation underlies a transcriptional memory of somatic cells in human iPS cells

Abstract

Human induced pluripotent stem (iPS) cells are remarkably similar to embryonic stem (ES) cells, but recent reports indicate that there may be important differences between them. We carried out a systematic comparison of human iPS cells generated from hepatocytes (representative of endoderm), skin fibroblasts (mesoderm) and melanocytes (ectoderm). All low-passage iPS cells analysed retain a transcriptional memory of the original cells. The persistent expression of somatic genes can be partially explained by incomplete promoter DNA methylation. This epigenetic mechanism underlies a robust form of memory that can be found in iPS cells generated by multiple laboratories using different methods, including RNA transfection. Incompletely silenced genes tend to be isolated from other genes that are repressed during reprogramming, indicating that recruitment of the silencing machinery may be inefficient at isolated genes. Knockdown of the incompletely reprogrammed gene C9orf64 (chromosome 9 open reading frame 64) reduces the efficiency of human iPS cell generation, indicating that somatic memory genes may be functionally relevant during reprogramming.

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Figure 1: Pluripotency validation for the derived Hep-iPS cells used for the microarray studies.
Figure 2: Multiple cell types undergo extensive transcriptional reprogramming to the human iPS cell state.
Figure 3: iPS cells retain a transcriptional memory of the original somatic cell.
Figure 4: DNA methylation can partially explain somatic gene expression in iPS cells.
Figure 5: Meta-analysis of DNA-methylation-associated transcriptional memory in independent data sets.
Figure 6: The somatic cell memory gene C9orf64 is required for efficient generation of iPS cells.
Figure 7: Proximity in the genome affects efficiency of gene silencing in iPS cells.
Figure 8: Model for the role of DNA methylation in reprogramming to the human iPS cell state.

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Acknowledgements

The authors wish to thank S. Fisher, O. Genbachev, A. Leavitt and B. Conklin for expert advice on culturing human ES cells, D. Subramanyam and R. Blelloch for the Adult Fibroblast-iPS 1 cell line, L. Ta, A. Williams and A. Holloway at the Gladstone Institutes, J. Bolen at the Mouse Pathology Core Facility for expert assistance, J. Utikal for technical advice, and J. Yang and A. Campain for sharing their meta-DEDS code. We thank members of the Santos laboratory, R. Blelloch, H. Willenbring, S. Fisher and M. Grskovic for helpful discussions and critical reading of the manuscript. Work in the Santos laboratory is supported by CIRM, JDRF, an NIH Director’s New Innovator Award and the Leona M. and Harry B. Helmsley Charitable Trust. Y.O. was partially supported by the UCSF Diabetes Center and a T32 grant from the NICHD to the UCSF Center for Reproductive Sciences. Work in M.H.’s laboratory was supported by grants from the JDRF and the Leona M. and Harry B. Helmsley Charitable Trust. T.G. was supported by the JDRF and the Leona M. and Harry B. Helmsley Charitable Trust. S.L.D. was partially supported by CIRM. J.S.S. was partially supported by the PhRMA Foundation.

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Contributions

Y.O., J.S.S. and M.R-S. conceived the project. J.M.P., K.H., P.D.M. and D.J.R. provided reagents. Z.Q. and J.Y. provided assistance with data analysis. C.H. and S.L.D. carried out the bisulphite sequencing analysis under supervision of J.F.C. T.G. carried out the targeted differentiation to endoderm analysis under supervision of M.H. J.S.S. carried out all of the bioinformatic analyses. Y.O., H.Q. and M.R-S. designed and Y.O. and H.Q. carried out all other experiments with technical assistance from L.B. Y.O, J.S.S. and M.R-S. wrote the manuscript with input from the other authors.

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Correspondence to Jun S. Song or Miguel Ramalho-Santos.

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Ohi, Y., Qin, H., Hong, C. et al. Incomplete DNA methylation underlies a transcriptional memory of somatic cells in human iPS cells. Nat Cell Biol 13, 541–549 (2011). https://doi.org/10.1038/ncb2239

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