Tracking epigenetic histone modifications in single cells using Fab-based live endogenous modification labeling
Open Access
- 14 May 2011
- journal article
- research article
- Published by Oxford University Press (OUP) in Nucleic Acids Research
- Vol. 39 (15), 6475-6488
- https://doi.org/10.1093/nar/gkr343
Abstract
Histone modifications play an important role in epigenetic gene regulation and genome integrity. It remains largely unknown, however, how these modifications dynamically change in individual cells. By using fluorescently labeled specific antigen binding fragments (Fabs), we have developed a general method to monitor the distribution and global level of endogenous histone H3 lysine modifications in living cells without disturbing cell growth and embryo development. Fabs produce distinct nuclear patterns that are characteristic of their target modifications. H3K27 trimethylation-specific Fabs, for example, are concentrated on inactive X chromosomes. As Fabs bind their targets transiently, the ratio of bound and free molecules depends on the target concentration, allowing us to measure changes in global modification levels. High-affinity Fabs are suitable for mouse embryo imaging, so we have used them to monitor H3K9 and H3K27 acetylation levels in mouse preimplantation embryos produced by in vitro fertilization and somatic cell nuclear transfer. The data suggest that a high level of H3K27 acetylation is important for normal embryo development. As Fab-based live endogenous modification labeling (FabLEM) is broadly useful for visualizing any modification, it should be a powerful tool for studying cell signaling and diagnosis in the future.Keywords
This publication has 39 references indexed in Scilit:
- Epigenetic reprogramming and development: a unique heterochromatin organization in the preimplantation mouse embryoBriefings in Functional Genomics, 2010
- Histone Deacetylase Inhibitors Stimulate Histone H3 Lysine 4 Methylation in Part Via Transcriptional Repression of Histone H3 Lysine 4 DemethylasesPublished by American Society for Pharmacology & Experimental Therapeutics (ASPET) ,2010
- Characterization of an antagonistic switch between histone H3 lysine 27 methylation and acetylation in the transcriptional regulation of Polycomb group target genesNucleic Acids Research, 2010
- Dissecting the binding mechanism of the linker histone in live cells: an integrated FRAP analysisThe EMBO Journal, 2010
- In Vivo Residue-specific Histone Methylation Dynamics*Published by Elsevier BV ,2010
- Visualizing histone modifications in living cells: spatiotemporal dynamics of H3 phosphorylation during interphaseThe Journal of cell biology, 2009
- G9a selectively represses a class of late-replicating genes at the nuclear peripheryProceedings of the National Academy of Sciences of the United States of America, 2009
- Fluorescent biosensors for real-time tracking of post-translational modification dynamicsCurrent Opinion in Chemical Biology, 2009
- Real-time imaging of histone H4 hyperacetylation in living cellsProceedings of the National Academy of Sciences of the United States of America, 2009
- CBP-mediated acetylation of histone H3 lysine 27 antagonizesDrosophilaPolycomb silencingDevelopment, 2009