Part of the book series:Methods in Molecular Biology ((MIMB,volume 463))
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Abstract
Epigenetic mechanisms lead to the stable regulation of gene expression without alteration of DNA and trigger initiation and/or maintenance of cell-type-specific transcriptional profiles. Indeed, modulation of chromatin structure and the global 3D organization of the genome and nuclear architecture participate in the precise control of transcription. Thus, dissection of these epigenetic mechanisms is essential for our understanding of gene regulation. In this chapter, we describe challenging combinations of immunofluorescence, and RNA and DNA fluorescent in situ hybridization and their application to our studies of a remarkable example of epigenetic control of gene expression in female mammals, the process of X chromosome inactivation.
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Fischle, W., Wang, Y., and Allis, C.D. (2003) Histone and chromatin cross-talk.Curr. Opin. Cell Biol.15, 172–183.
Dillon, N. (2006) Gene regulation and large-scale chromatin organization in the nucleus.Chrom. Res.14, 117–126.
Lawrence, J.B. and Singer, R.H. (1985) Quantitative analysis of in situ hybridization methods for the detection of actin gene expression.Nucleic Acids Res.13, 1777–1799.
Foster, H.A. and Bridger, J.M. (2005) The genome and the nucleus: a marriage made by evolution. Genome organization and nuclear structure.Chromosoma114, 212–219.
Baxter, J., Merkenschlager, M., and Fisher, A.G. (2002) Nuclear organization and gene expression.Curr. Opin. Cell Biol.14, 372–376.
Chambeyron, S. and Bickmore, W.A. (2004) Does looping and clustering in the nucleus regulate gene expression?Curr. Opin. Cell Biol.16, 256–262.
Nightingale, K.P., O’Neill, L.P., and Turner, B.M. (2006) Histone modifications: Signalling receptors and potential elements of a heritable epigenetic code.Curr. Opin. Gen. Dev.16, 125–136.
Mattick, J.S. and Makunin, I.V. (2006) Non-coding RNA.Hum. Mol. Genet.15, 17–29.
Heard, E. and Disteche, C.M. (2006) Dosage compensation in mammals: fine-tuning the expression of the X chromosome.Genes Dev.20, 1848–1867.
Chaumeil, J., Le Baccon, P., Wutz, A., and Heard, E. (2006) A novel role for Xist RNA in the formation of a repressive nuclear compartment into which genes are recruited when silenced.Genes Dev.20, 2223–2237.
Chaumeil, J., Okamoto, I., Guggiari, M., and Heard, E. (2002) Integrated kinetics of X-chromosome inactivation in differentiating embryonic stem cells.Cytogen. Gen. Res.99, 75–84.
Chaumeil, J., Okamoto, I., and Heard, E. (2004) X-inactivation in mouse embryonic stem cells: Analysis of histone modifications and transcriptional activity using immunofluores-cence and FISH.Methods Enzymol.376, 405–419.
Chaumeil, J. (2005) Immunofluorescence—fluorescent in situ hybridization. Protocol for the Epigenome Network of Excellence (http://www.epigenome-noe.net).
Spector, D.L., Goldman, R.D., and Leinwand, L.A. (1998)Cells: a laboratory manual. Cold Spring Harbor Laboratory Press: Cold Spring Harbor, N Y.
Acknowledgments
We thank the EU Network of Excellence “The Epigenome” for permission to republish these procedures. Funding sources for this work were the Human Frontiers Science Program, EU FP6 Integrated Project HEROIC (LSHG-CT-2005-018883), the Schlumberger Foundation for Research, and the EU FP6 Network of Excellence “The Epigenome” (LSHG-CT-2004-503433). JC was funded by the French “Ligue Nationale contre le Cancer” and the HEROIC IP; SA was funded by the French “Centre National de la Recherche Scientifique”; JCC was funded by the HEROIC IP.
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Authors and Affiliations
Mammalian Developmental Epigenetics Group, CNRS UMR 218, Curie Institute, Paris, France
Julie Chaumeil, Sandrine Augui, Jennifer C. Chow & Edith Heard
- Julie Chaumeil
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- Sandrine Augui
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- Jennifer C. Chow
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- Edith Heard
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Hôtel-Dieu Hospital, Laval University Cancer Research Centre, Québec, Canada
Ronald Hancock
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© 2008 Humana Press
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Chaumeil, J., Augui, S., Chow, J.C., Heard, E. (2008). Combined Immunofluorescence, RNA Fluorescent In Situ Hybridization, and DNA Fluorescent In Situ Hybridization to Study Chromatin Changes, Transcriptional Activity, Nuclear Organization, and X-Chromosome Inactivation. In: Hancock, R. (eds) The Nucleus. Methods in Molecular Biology, vol 463. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-59745-406-3_18
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