Auxin-inducible degron 2 system deciphers functions of CTCF domains intranscriptional regulation.

Hyle J. et al.

BACKGROUND: CTCF is a well-established chromatin architectural protein that also plays various roles in transcriptional regulation. While CTCF biology has been extensively studied, how the domains of CTCF function to regulate transcription remains unknown. Additionally, the original auxin-inducible degron 1 (AID1) system has limitations in investigating the function of CTCF. RESULTS: We employ an improved auxin-inducible degron technology, AID2, to facilitate the study of acute depletion of CTCF while overcoming the limitations of the previous AID system. As previously observed through the AID1 system and steady-state RNA analysis, the new AID2 system combined with SLAM-seq confirms that CTCF depletion leads to modest nascent and steady-state transcript changes. A CTCF domain sgRNA library screening identifies the zinc finger (ZF) domain as the region within CTCF with the most functional relevance, including ZFs 1 and 10. Removal of ZFs 1 and 10 reveals genomic regions that independently require these ZFs for DNA binding and transcriptional regulation. Notably, loci regulated by either ZF1 or ZF10 exhibit unique CTCF binding motifs specific to each ZF. CONCLUSIONS: By extensively comparing the AID1 and AID2 systems for CTCF degradation in SEM cells, we confirm that AID2 degradation is superior for achieving miniAID-tagged protein degradation without the limitations of the AID1 system. The model we create that combines AID2 depletion of CTCF with exogenous overexpression of CTCF mutants allows us to demonstrate how peripheral ZFs intricately orchestrate transcriptional regulation in a cellular context for the first time.


Share this article

January, 2023


Products used in this publication

  • cut and tag antibody icon
    CTCF Antibody


  • APHL 2024
    Milwaukee, Wisconsin, USA
    May 6-May 9, 2024
 See all events


 See all news

The European Regional Development Fund and Wallonia are investing in your future.

Extension of industrial buildings and new laboratories.

       Site map   |   Contact us   |   Conditions of sales   |   Conditions of purchase   |   Privacy policy   |   Diagenode Diagnostics