Contributor:

Pękowska, Aleksandra (1982- ) : Supervisor

Publisher:

Instytut Biologii Doświadczalnej im. Marcelego Nenckiego PAN

Place of publishing:

Warszawa

Description:

173 pages : illustrations ; 30 cm ; Bibliography ; Summary in English

Degree name:

PhD in Biological Sciences

Degree discipline :

Biological Sciences

Degree grantor:

Nencki Institute of Experimental Biology PAS ; degree obtained: 17.04.2026

Type of object:

Thesis

Abstract:

CTCF is a conserved DNA-binding protein that plays a key role in regulating the three- dimensional architecture of the genome. It defines the boundaries of topologically associated domains (TADs) and controls interactions between promoters and enhancers. In mammalian cells, the positions of CTCF-binding sites (CBS) have been shown to remain essentially unchanged during embryonic stem (ES) cell to neural stem (NS) cell differentiation, despite an increase in the number and stability of chromatin loops. Changes in chromatin architecture involving CTCF may influence its role in regulating gene expression. However, it remains unclear whether, and how, this regulatory function changes during development. This doctoral thesis aimed to address this question and assess the functions of CTCF during the differentiation of ES cells into NS cells. To investigate this, a CTCF degron system enabling precise, inducible protein depletion was used, followed by transcriptome and regulome profiling in both cellular states. CTCF loss altered gene expression, but did not directly affect regulatory element activity. Genes downregulated upon CTCF depletion were highly expressed and contained CTCF binding at their promoters, whereas upregulated genes showed low basal expression and lacked promoter-bound CTCF. These findings support a model in which promoter- bound CTCF facilitates transcription by mediating contacts with distal enhancers, while intergenic CTCF acts as an insulator. The experiments outlined in this dissertation revealed that, despite stable DNA binding, CTCF regulates distinct gene sets in ES and NS cells, indicating cell-type- specific functions of this factor in development. This was further validated by genome editing at the Aldh1a3 locus. To unveil what underlies the changes in CTCF functions in development, proteomic analysis was performed independently in the lab. These data revealed enhanced interactions between CTCF and RNA-binding proteins (RBP) in the NS compared to the ES cells. The cells derived in this work help identify long non-coding RNA (lncRNA) Pantr1 as a mediator of the gain of CTCF-RBP interactions upon the ES-to-NS transition. In addition, a visualisation system was developed to monitor selected genomic loci in living cells, which has the potential, in the future, to enable real-time tracking of chromatin loop anchor dynamics. In summary, although the localisation of CTCF binding sites remains stable during differentiation, its transcriptional regulatory function is dynamic and highly context-dependent.

Detailed Resource Type:

PhD Dissertations

Resource Identifier:

oai:rcin.org.pl:262616

Source:

IBD PAN, call no. 20927

Language:

pol

Language of abstract:

eng

Rights:

Rights Reserved - Free Access

Terms of use:

Copyright-protected material. May be used within the limits of statutory user freedoms

Copyright holder:

Publication made available with the written permission of the author

Digitizing institution:

Nencki Institute of Experimental Biology of the Polish Academy of Sciences

Original in:

Library of the Nencki Institute of Experimental Biology PAS

Access:

Open