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Embryonic stem cell–based mapping of developmental transcriptional programs

Nature Methods volume 8, pages 1056–1058 (2011)Cite this article

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Abstract

The study of developmentally regulated transcription factors by chromatin immunoprecipitation and deep sequencing (ChIP-seq) faces two major obstacles: availability of ChIP-grade antibodies and access to sufficient number of cells. We describe versatile genome-wide analysis of transcription-factor binding sites by combining directed differentiation of embryonic stem cells and inducible expression of tagged proteins. We demonstrate its utility by mapping DNA-binding sites of transcription factors involved in motor neuron specification.

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Figure 1: Generation of inducible cell lines.
Figure 2: Native- and tagged-protein ChIP comparisons.

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Gene Expression Omnibus

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Acknowledgements

E.O.M. is funded as the David and Sylvia Lieb Fellow of the Damon Runyon Cancer Research Foundation (DRG-1937-07). We thank R. Sherwood (Harvard University) for sharing unpublished observations and T Jessell (Columbia University) for anti-Hoxc4 antibody. Personnel and work were supported by US National Institutes of Health grant P01 NS055923 (D.K.G., R.A.Y. and H.W.), R01 NS058502 (H.W.), R01HL081186 (M.K.) and Helmsley Stem Cell Starter grant (H.W.).

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Authors and Affiliations

  1. Departments of Pathology, Neurology and Neuroscience, Center for Motor Neuron Biology and Disease and Columbia Stem Cell Initiative, Columbia University Medical Center, New York, New York, USA

    Esteban O Mazzoni, Carolyn A Morrison, George Mountoufaris, Michael Closser & Hynek Wichterle

  2. Computer Science and Artificial Intelligence Laboratory, Massachusetts Institute of Technology, Cambridge, Massachusetts, USA

    Shaun Mahony & David K Gifford

  3. Lillehei Heart Institute and Department of Pediatrics, University of Minnesota, Minneapolis, Minnesota, USA

    Michelina Iacovino & Michael Kyba

  4. Department of Biology, Massachusetts Institute of Technology, Cambridge, Massachusetts, USA

    Warren A Whyte & Richard A Young

  5. Whitehead Institute for Biomedical Research, Cambridge, Massachusetts, USA

    Warren A Whyte & Richard A Young

Authors
  1. Esteban O Mazzoni
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  2. Shaun Mahony
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  3. Michelina Iacovino
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  4. Carolyn A Morrison
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  5. George Mountoufaris
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  6. Michael Closser
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  7. Warren A Whyte
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  8. Richard A Young
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  9. Michael Kyba
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  10. David K Gifford
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  11. Hynek Wichterle
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Contributions

E.O.M. and G.M. generated the transcription factor inducible lines; E.O.M. performed phenotypic analysis of the derived lines. E.O.M., W.A.W. and C.A.M. performed ChIP experiments. M.I. and M.K. developed the ICE cell lines and vectors. E.O.M. performed expression analysis. E.O.M. and M.C. performed the western blot and protein-binding to immobilized DNA. S.M. analyzed ChIP-seq data. E.O.M., R.A.Y., D.K.G .and H.W. designed the experiments. E.O.M., S.M. and H.W. wrote the manuscript; D.K.G. revised the manuscript.

Corresponding author

Correspondence to Hynek Wichterle.

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The authors declare no competing financial interests.

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Supplementary Figures 1–3 and Supplementary Table 1 (PDF 1439 kb)

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Mazzoni, E., Mahony, S., Iacovino, M. et al. Embryonic stem cell–based mapping of developmental transcriptional programs. Nat Methods 8, 1056–1058 (2011). https://doi.org/10.1038/nmeth.1775

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