In:
Journal of Cell Science, The Company of Biologists
Abstract:
Chromatin modification is traditionally queried by biochemical assays that provide average measurements of static events since the analysis requires components from many cells. Microscopy can visualize single cells, but the cell body and organelles can hamper staining and visualization of the nucleus. Normally, chromatin is visualized by immunostaining a fixed sample or by expressing exogenous fluorescently tagged proteins in a live cell. Alternative microscopy tools to observe changes of endogenous chromatin in real-time are needed. Here, we isolated transcriptionally competent nuclei from cells and used antibody staining without fixation to visualize changes of endogenous chromatin. This method allows the real-time addition of drugs and fluorescent probes to one or more nuclei while under microscopic observation. A high-resolution map of eleven endogenous nuclear markers of the histone code, transcription machinery, and architecture was obtained in transcriptionally active nuclei with confocal and structured illumination microscopy. We detected changes in chromatin modification and localization at the single-nucleus level after inhibition of histone deacetylation. Applications in the study of RNA transcription, viral protein function and nuclear architecture are presented.
Type of Medium:
Online Resource
ISSN:
1477-9137
,
0021-9533
Language:
English
Publisher:
The Company of Biologists
Publication Date:
2017
detail.hit.zdb_id:
219171-4
detail.hit.zdb_id:
1483099-1
SSG:
12
Permalink
