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Related Concept Videos

Chromatin Position Affects Gene Expression02:35

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Chromatin is the massive complex of DNA and proteins packaged inside the nucleus. The complexity of chromatin folding and how it is packaged inside the nucleus greatly influences  access to genetic information. Generally, the nucleus' periphery is considered transcriptionally repressive, while the cell's interior is considered a transcriptionally active area. 
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Cis-regulatory sequences are short fragments of non-coding DNA that are present on the same chromosomes as the genes that they regulate. These fragments serve as binding sites for transcriptional regulators, proteins that are responsible for controlling gene transcription and differential gene expression across cell types in eukaryotes. Cis-regulatory sequences can be close to the gene of interest or thousands of bases away in the DNA sequence; however, those sequences that are further away are...
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The histone proteins in the nucleosomes are post-translationally modified (PTM) to increase or decrease access to DNA. The commonly observed PTMs are methylation, acetylation, phosphorylation, and ubiquitination of lysine amino acids in the histone H3 tail region. These histone modifications have specific meaning for the cell. Hence, they are called "histone code". The protein complex involved in histone modification is termed as "reader-writer" complex.
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Epigenetics is the study of inherited changes in a cell's phenotype without changing the DNA sequences. It provides a form of memory for the differential gene expression pattern to maintain cell lineage, position-effect variegation, dosage compensation, and maintenance of chromatin structures such as telomeres and centromeres. For example, the structure and location of the centromere on chromosomes are epigenetically inherited. Its functionality is not dictated or ensured by the underlying...
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Multicellular organisms contain a variety of structurally and functionally distinct cell types, but the DNA in all the cells originated from the same parent cells. The differences in the cells can be attributed to the differential gene expression. Liver cells, whose functions include detoxification of blood, production of bile to metabolize fats, and synthesis of proteins essential for metabolism, must express a specific set of genes to perform their functions. Gene expression also varies with...
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Each human somatic cell contains 6 billion base-pairs of DNA. Each base-pair is 0.34 nm long, which means that each diploid cell contains a staggering 2 meters of DNA. How is such a long DNA strand packed inside a nucleus measuring only 10 - 20 microns in diameter? 
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Automated in situ chromatin profiling efficiently resolves cell types and gene regulatory programs.

Derek H Janssens1, Steven J Wu1,2, Jay F Sarthy1,3

  • 1Basic Sciences Division, Fred Hutchinson Cancer Research Center, 1100 N. Fairview Ave, Seattle, WA, 98109, USA.

Epigenetics & Chromatin
|December 23, 2018
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Automated CUT&RUN efficiently maps chromatin landscapes and protein-DNA interactions. This cost-effective method distinguishes cell types and tumor subtypes, advancing gene regulation studies.

Keywords:
CUT&RUNChromatin regulatorsHistone modificationsTranscription factors

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Area of Science:

  • Genomics
  • Molecular Biology
  • Epigenetics

Background:

  • Eukaryotic gene regulation is complex due to intricate protein-DNA interactions within the chromatin landscape.
  • Current methods for characterizing these interactions are often inefficient.
  • CUT&RUN (Cleavage Under Targets and Release) is a sensitive, high-resolution in situ method for profiling DNA-binding proteins, histones, and chromatin-modifying proteins.

Purpose of the Study:

  • To describe an automated CUT&RUN platform.
  • To apply automated CUT&RUN for characterizing human cell chromatin landscapes.
  • To evaluate the utility of automated CUT&RUN for profiling tumor samples.

Main Methods:

  • Development and implementation of an automated CUT&RUN workflow.
  • Application of automated CUT&RUN to profile histone modifications in human cells.
  • Analysis of automated CUT&RUN data to identify cell-type-specific regulatory elements.
  • Profiling of frozen pediatric glioma xenograft samples using automated CUT&RUN.

Main Results:

  • Automated CUT&RUN profiles of histone modifications accurately demarcate active and repressed chromatin regions.
  • A continuous metric was developed to identify cell-type-specific promoter and enhancer activities.
  • Automated CUT&RUN successfully distinguished between two pediatric glioma xenografts based on their distinct gene expression programs.
  • The method demonstrated high sensitivity and resolution in profiling chromatin landscapes.

Conclusions:

  • The automated CUT&RUN workflow is easy to implement and cost-effective.
  • Automated CUT&RUN is a valuable tool for high-throughput characterization of diverse cell types.
  • This method facilitates the analysis of patient samples, including tumors, for subtype-specific molecular signatures.