Generation of germline-competent induced pluripotent stem cells (original) (raw)
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- Published: 06 June 2007
Nature volume 448, pages 313–317 (2007)Cite this article
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Abstract
We have previously shown that pluripotent stem cells can be induced from mouse fibroblasts by retroviral introduction of Oct3/4 (also called Pou5f1), Sox2, c-Myc and Klf4, and subsequent selection for Fbx15 (also called Fbxo15) expression. These induced pluripotent stem (iPS) cells (hereafter called Fbx15 iPS cells) are similar to embryonic stem (ES) cells in morphology, proliferation and teratoma formation; however, they are different with regards to gene expression and DNA methylation patterns, and fail to produce adult chimaeras. Here we show that selection for Nanog expression results in germline-competent iPS cells with increased ES-cell-like gene expression and DNA methylation patterns compared with Fbx15 iPS cells. The four transgenes (Oct3/4, Sox2, c-myc and Klf4) were strongly silenced in Nanog iPS cells. We obtained adult chimaeras from seven Nanog iPS cell clones, with one clone being transmitted through the germ line to the next generation. Approximately 20% of the offspring developed tumours attributable to reactivation of the c-myc transgene. Thus, iPS cells competent for germline chimaeras can be obtained from fibroblasts, but retroviral introduction of c-Myc should be avoided for clinical application.
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Acknowledgements
We thank K. Takahashi, M. Nakagawa and T. Aoi for scientific discussion; M. Maeda for histological analyses; M. Narita, J. Iida, H. Miyachi and S. Kitano for technical assistance; and R. Kato, R. Iyama and Y. Ohuchi for administrative assistance. We also thank T. Kitamura for Plat-E cells and pMXs retroviral vectors, and R. Farese for RF8 ES cells. This study was supported in part by a grant from the Uehara Memorial Foundation, the Program for Promotion of Fundamental Studies in Health Sciences of NIBIO, a grant from the Leading Project of MEXT, and Grants-in-Aid for Scientific Research of JSPS and MEXT (to S.Y.). K.O. is a JSPS research fellow. The microarray data are deposited in GEO under accession number GSE7841.
Author Contributions K.O. conducted most of the experiments in this study. T.I. performed manipulation of mouse embryos to generate Nanog-GFP transgenic mice. T.I. also maintained the mouse lines. S.Y. designed and supervised the study, and prepared the manuscript. S.Y. also performed computer analyses of DNA microarray data.
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Authors and Affiliations
- Department of Stem Cell Biology, Institute for Frontier Medical Sciences, Kyoto University, Kyoto 606-8507, Japan,
Keisuke Okita, Tomoko Ichisaka & Shinya Yamanaka - CREST, Japan Science and Technology Agency, Kawaguchi 332-0012, Japan,
Tomoko Ichisaka & Shinya Yamanaka
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- Keisuke Okita
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Corresponding author
Correspondence toShinya Yamanaka.
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Competing interests
The microarray data are deposited in GEO under accession number GSE7841. Reprints and permissions information is available at www.nature.com/reprints. The authors declare no competing financial interests.
Supplementary information
Supplementary Information
This file contains Supplementary Figures 1-13 with Legends and Supplementary Table 1, which show detailed characterization of Nanog-iPS cells. (PDF 1076 kb)
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Okita, K., Ichisaka, T. & Yamanaka, S. Generation of germline-competent induced pluripotent stem cells.Nature 448, 313–317 (2007). https://doi.org/10.1038/nature05934
- Received: 06 February 2007
- Accepted: 22 May 2007
- Published: 06 June 2007
- Issue Date: 19 July 2007
- DOI: https://doi.org/10.1038/nature05934
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Editorial Summary
Stem cells with potential
The search for new ways of coaxing readily available cells to show the pluripotent activity of embryonic stem cells — the potential to differentiate into virtually any cell type — continues. The stakes are high, since if it can be achieved safely for human cells, cell transplantation therapy, even patient-specific therapy, will have come a step closer. Two groups now report an important advance in this direction: the creation of pluripotent stem cells from mouse fibroblasts. The epigenetic reprogramming requires the expression of four transcription factors, Oct3/4, Sox2, c-Myc and Klf4. The resulting cells resemble embryonic stem cells in both biological potency and epigenetic state.