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Related Concept Videos

Heterochromatin02:38

Heterochromatin

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The extent of chromatin compaction can be studied by staining chromatin using specific DNA binding dyes. Under the microscope, the dense-compacted regions that take up more dye are called heterochromatin. Heterochromatin is further classified into two forms – constitutive heterochromatin and facultative heterochromatin.
Constitutive heterochromatin: It is a highly compact region of chromatin that is mostly concentrated in the centromere and telomere. Unlike euchromatin, the amino acid at...
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Histone variants are the histone proteins with structural and sequence variations. These variants may be regarded as “mutant” forms that replace their canonical histone counterparts in the nucleosomes. Specific post-translational modifications on the histone variants enable further chromatin complexity and regulate tissue-specific gene expression. The most common histone variants are from histone H2A, H2B, and linker histone H1 families. However, several variants of histone H3...
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As cells progress into mitosis, the nuclear envelope breaks down, and the condensed chromosomes are exposed to the array of bipolar microtubules of the mitotic spindle. The kinetochore, a large, disc-shaped protein complex, is present at the centromere region of the sister chromatids and acts as a binding site for the microtubules.  Usually, the plus-end of a single microtubule is embedded within the kinetochore. However, some kinetochores first establish lateral contact with the side-wall...
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A functional eukaryotic chromosome must contain three elements: a centromere, telomeres, and numerous origins of replication.
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Chromatin modification alters gene expression; therefore, scientists can add histone-modifying enzymes, histone variants, and chromatin remodeling complexes to somatic cells to aid reprogramming into pluripotent stem (iPS) cells.
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Hi-C: A Method to Study the Three-dimensional Architecture of Genomes.
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Identifying multi-locus chromatin contacts in human cells using tethered multiple 3C.

Ferhat Ay1, Thanh H Vu2, Michael J Zeitz3

  • 1Department of Genome Sciences, University of Washington, Seattle, 98195, WA, USA. ferhatay@uw.edu.

BMC Genomics
|April 19, 2015
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Summary

A new Tethered Multiple Chromatin Conformation Capture (TM3C) assay maps genome-wide 3D chromatin contacts. This method identifies simultaneous multi-locus interactions, revealing how open chromatin regions cluster in near-haploid cells.

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Area of Science:

  • Genomics
  • Molecular Biology
  • Epigenetics

Background:

  • Advanced chromatin conformation capture (3C) methods profile genome-wide chromatin contacts in 3D.
  • Chromatin folding in complex eukaryotes correlates with gene expression and DNA replication.

Purpose of the Study:

  • To introduce Tethered Multiple Chromatin Conformation Capture (TM3C), a novel assay for genome-wide chromatin contact mapping.
  • To characterize the genome architecture of human cell lines using TM3C, including pairwise and multi-way contacts.

Main Methods:

  • Tethered Multiple Chromatin Conformation Capture (TM3C) protocol involving restriction enzyme digestion and religation of fragments on agarose gel beads.
  • Paired-end sequencing for mapping genome-wide chromatin contacts.
  • Analysis of contact frequency maps, including scaling with genomic distance, chromosomal compartments, and topological domains.

Main Results:

  • TM3C successfully mapped genome-wide chromatin contacts in KBM7 (near-haploid) and NHEK (diploid) human cell lines.
  • The assay identified simultaneous contacts among multiple loci, including cell type-specific contacts, ploidy shifts, and translocations like the Philadelphia chromosome.
  • Genome-wide analysis revealed preferential linking of open chromatin regions and regions with high DNase hypersensitive sites (DHSs).
  • 3D models of near-haploid KBM7 cells showed clustering of small and large chromosomes.

Conclusions:

  • TM3C is a straightforward method for determining genome architecture and identifying simultaneous contacts among three or four loci.
  • TM3C application to a near-haploid cell line elucidated large-scale chromosomal organization and multi-way chromatin contacts that link open chromatin regions.