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Updated: Jun 13, 2025

Immunofluorescence Analysis of Endogenous and Exogenous Centromere-kinetochore Proteins
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CTCF maintains pericentromere function and mitotic fidelity.

Erin Walsh1, Andrew D Stephens1,2

  • 1Biology department, University of Massachusetts Amherst, Amherst, MA 01003, USA.

Biorxiv : the Preprint Server for Biology
|June 12, 2025
PubMed
Summary
This summary is machine-generated.

CTCF protein is crucial for cell division (mitosis). Degrading CTCF disrupts chromosome alignment and nuclear shape, revealing its role in maintaining pericentromere function and successful mitosis.

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Area of Science:

  • Cell Biology
  • Genetics
  • Molecular Biology

Background:

  • Mitosis requires accurate genome segregation.
  • CTCF (CCCTC-binding factor) is a known interphase chromatin regulator.
  • CTCF's role in mitosis was previously unknown, though its depletion caused mitotic failure.

Purpose of the Study:

  • To elucidate the specific role of CTCF in mitosis.
  • To investigate the molecular mechanisms underlying CTCF-dependent mitotic regulation.

Main Methods:

  • Utilized a CRISPR-Cas9 auxin-inducible degron system for rapid CTCF depletion in human cells.
  • Performed immunofluorescence microscopy to analyze mitotic spindle organization and chromosome alignment.
  • Assessed post-mitotic nuclear morphology.

Main Results:

  • CTCF degradation led to increased mitotic failures and abnormal post-mitotic nuclear shapes.
  • Kinetochore recruitment of CENP-E remained intact, and polar chromosomes were infrequent.
  • CTCF depletion caused wider intercentromere distances and a disorganized metaphase plate, mimicking cohesin loss.
  • Disruption of pericentromere function was observed.

Conclusions:

  • CTCF is essential for maintaining pericentromere integrity and function during mitosis.
  • CTCF plays a critical role in ensuring accurate chromosome segregation and proper post-mitotic nuclear formation.
  • CTCF acts as a key factor in successful mitosis and the maintenance of nuclear shape.