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Auxin-inducible degron 2 system deciphers functions of CTCF domains in transcriptional regulation

ABSTRACT: This SuperSeries is composed of the SubSeries listed below.

ORGANISM(S): Homo sapiens

PROVIDER: GSE205218 | GEO | 2022/12/09

REPOSITORIES: GEO

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

Project description:BackgroundCTCF 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.ResultsWe 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.ConclusionsBy 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.

| S-EPMC9878928 | biostudies-literature

Auxin-inducible degron 2 system deciphers functions of CTCF domains in transcriptional regulation [HiChIP]

Project description: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 to investigate 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 transcripts 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.

2022-12-09 | GSE201911 | GEO

Auxin-inducible degron 2 system deciphers functions of CTCF domains in transcriptional regulation [RNA-Seq]

2022-12-09 | GSE205311 | GEO

Auxin-inducible degron 2 system deciphers functions of CTCF domains in transcriptional regulation [SLAM-seq]

2022-12-09 | GSE205184 | GEO

Advanced auxin-inducible degron system deciphers unique functions of CTCF in transcriptional regulation [ChIP-seq]

Project description:ChIPseq was performed on cells depleted for CTCF by AID2 auxin degradation to examine CTCF DNA binding affinity. ChIP seq was also performed on cells depleted for endogenous CTCF and expressing HA tagged WT CTCF and CTCF mutants (deltaZF1, deltaZF10 and deltaRBR) to examine how functional deletions affect CTCF DNA binding affinity.

2022-12-28 | GSE205408 | GEO

Auxin-inducible degron 2 system deciphers functions of CTCF domains in transcriptional regulation

Project description:Auxin-inducible degron 2 system deciphers functions of CTCF domains in transcriptional regulation

| PRJNA844097 | ENA

Auxin-inducible degron 2 system deciphers functions of CTCF domains in transcriptional regulation [HiChIP]

Project description:Auxin-inducible degron 2 system deciphers functions of CTCF domains in transcriptional regulation [HiChIP]

| PRJNA833451 | ENA

Auxin-inducible degron 2 system deciphers functions of CTCF domains in transcriptional regulation [RNA-Seq]

Project description:Auxin-inducible degron 2 system deciphers functions of CTCF domains in transcriptional regulation [RNA-Seq]

| PRJNA844363 | ENA

Auxin-inducible degron 2 system deciphers functions of CTCF domains in transcriptional regulation [SLAM-seq]

Project description:Auxin-inducible degron 2 system deciphers functions of CTCF domains in transcriptional regulation [SLAM-seq]