Opposing Effects of Cohesin and Transcription on CTCF Organization Revealed by Super-resolution Imaging

Bo Gu, Colin J. Comerci, Dannielle G. McCarthy, Saumya Saurabh, W. E. Moerner, Joanna Wysocka

Research output: Contribution to journalArticlepeer-review

Abstract

CCCTC-binding factor (CTCF) and cohesin play critical roles in organizing mammalian genomes into topologically associating domains (TADs). Here, by combining genetic engineering with quantitative super-resolution stimulated emission depletion (STED) microscopy, we demonstrate that in living cells, CTCF forms clusters typically containing 2–8 molecules. A fraction of CTCF clusters, enriched for those with ≥3 molecules, are coupled with cohesin complexes with a characteristic physical distance suggestive of a defined molecular interaction. Acute degradation of the cohesin unloader WAPL or transcriptional inhibition (TI) result in increased CTCF clustering. Furthermore, the effect of TI on CTCF clusters is alleviated by the acute loss of the cohesin subunit SMC3. Our study provides quantitative characterization of CTCF clusters in living cells, uncovers the opposing effects of cohesin and transcription on CTCF clustering, and highlights the power of quantitative super-resolution microscopy as a tool to bridge the gap between biochemical and genomic methodologies in chromatin research.

Original languageEnglish (US)
Pages (from-to)699-711.e7
JournalMolecular Cell
Volume80
Issue number4
DOIs
StatePublished - Nov 19 2020

Keywords

  • chromatin
  • cohesin
  • CTCF
  • Pol II
  • quantitative analysis
  • super-resolution imaging
  • topological associated domain
  • transcription

ASJC Scopus subject areas

  • Molecular Biology
  • Cell Biology

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