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G-quadruplexes associated with R-loops promote CTCF binding.

Phillip Wulfridge1, Qingqing Yan1, Nathaniel Rell1

  • 1Gene expression and Regulation program, The Wistar Institute, Philadelphia, PA 19104, USA; Epigenetics Institute, University of Pennsylvania, Philadelphia, PA 19104, USA.

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|August 8, 2023
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Summary

R-loops and G-quadruplex (G4) structures facilitate CTCF binding to DNA, influencing genome architecture and gene expression. This discovery reveals how chromatin structure regulates CTCF occupancy in cells.

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CTCFG-quadruplexR-loopsgenome organization

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Area of Science:

  • Genomics
  • Molecular Biology
  • Epigenetics

Background:

  • CTCF (CCCTC-binding factor) is a key regulator of genome architecture and gene expression.
  • CTCF binding is influenced by DNA sequence motifs and modifications, but not all potential sites are occupied.
  • The role of chromatin structure in regulating CTCF occupancy remains incompletely understood.

Purpose of the Study:

  • To investigate the role of R-loops and G-quadruplex (G4) structures in regulating CTCF binding.
  • To determine if chromatin structures can influence CTCF occupancy at specific genomic sites.

Main Methods:

  • Co-localization studies of R-loops, G4s, and CTCF in mouse embryonic stem cells.
  • In vitro assays to assess CTCF binding to DNA motifs in the presence of R-loops and G4s.
  • In vivo experiments involving R-loop attenuation and G4 motif deletion/stabilization.

Main Results:

  • R-loops and G4s co-localize with CTCF at numerous genomic regions.
  • R-loops and G4s promote CTCF binding to its DNA motif in vitro.
  • R-loop attenuation decreases CTCF binding in vivo.
  • G4 motif deletion reduces CTCF binding and alters gene expression.
  • G4 stabilization increases CTCF occupancy and affects chromatin organization.

Conclusions:

  • R-loops facilitate CTCF binding by promoting the formation of associated G4 structures.
  • G4 structures play a pivotal role in reinforcing long-range genome interactions via CTCF.
  • Chromatin structural elements like R-loops and G4s are critical regulators of CTCF occupancy and function.