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Formaldehyde-assisted Isolation of Regulatory Elements to Measure Chromatin Accessibility in Mammalian Cells
Published on: April 2, 2018
Assaying chromatin structure and remodeling by restriction enzyme accessibility
Kevin W Trotter1, Trevor K Archer
1Laboratory of Molecular Carcinogenesis, National Institute of Environmental Health Sciences, National Institutes of Health, Research Triangle Park, NC, USA.
Methods in Molecular Biology (Clifton, N.J.)
|December 21, 2011
Summary
Chromatin remodeling complexes overcome DNA packaging barriers to enable nuclear processes. This study details a restriction endonuclease assay to quantify chromatin changes during remodeling, exemplified by BRG1 complex activity.
Area of Science:
- Molecular Biology
- Epigenetics
- Genomics
Background:
- Eukaryotic DNA is packaged into nucleosomes, forming chromatin, which acts as a barrier to essential nuclear processes like transcription and DNA replication.
- Chromatin remodeling complexes enzymatically alter chromatin structure, facilitating access for regulatory factors to target genes.
- Understanding chromatin dynamics is crucial for deciphering gene regulation and cellular functions.
Purpose of the Study:
- To present a detailed methodology for analyzing chromatin architecture and remodeling.
- To introduce the restriction endonuclease hypersensitivity assay for characterizing nucleosome remodeling.
- To demonstrate the application of this assay in studying specific chromatin remodeling events.
Main Methods:
- Utilizing restriction endonucleases to probe DNA accessibility within chromatin.
- Employing the restriction endonuclease hypersensitivity assay to detect changes in nucleosome positioning and structure.
- Analyzing the BRG1 complex-dependent chromatin remodeling of the mouse mammary tumor virus promoter as an experimental model.
Main Results:
- The restriction endonuclease hypersensitivity assay effectively characterizes alterations in chromatin structure.
- The methodology allows for the quantification of nucleosome remodeling events in response to regulatory signals.
- Specific changes in chromatin architecture mediated by the BRG1 complex were identified at the target promoter.
Conclusions:
- The restriction endonuclease hypersensitivity assay is a valuable tool for studying chromatin remodeling.
- This method provides quantitative insights into how chromatin structure is dynamically regulated.
- The findings contribute to a deeper understanding of gene regulation mechanisms involving chromatin dynamics.
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