Dependency of 3D chromatin folding on RNA polymerase II at different cell cycle stages

by Nadine Übelmesser
Doctoral thesis
Date of Examination:2023-06-29
Date of issue:2023-07-13
Advisor:Prof. Dr. Argyris Papantonis
Referee:Prof. Dr. Argyris Papantonis
Referee:Dr. Ufuk Günesdogan
Persistent Address: http://dx.doi.org/10.53846/goediss-9998

 

 

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Abstract

English

In human cells, the genetic information is organised in a multilayer fashion. During the cell cycle, chromatin undergoes dramatic structural changes. For proper inheritance, chromatin is compacted to chromosomes in mitosis, followed by reestablishment of the chromatin structure in G1 phase. Advances in technologies have provided insight into the fundamental question what shapes the genome architecture. By now, it is generally accepted that the 3D organisation is developed in a hierarchical manner, which is conserved between species. Moreover, several studies proposed that the 3D organisation could influence transcription. Despite an emerging picture on the 3D organisation, a structural contribution of RNA polymerase II (RNAPII) remains unclear. This study aims to contribute to this growing area of research by exploring the role of RNAPII in genome organization. I combined methods of molecular biology, transcriptomics, and genomics with the distinct auxin-inducible degradation of the largest RNAPII subunit RPB1 to elucidate its role in spatiotemporal chromatin structure regulation. In Chapter 1, I show that RNAP II is an important regulator in reestablishing the spatial chromatin structure at the mitosis to G1 transition. On a larger scale RNAPII depletion in G1-sorted cells erases domain structures, weakens local insulation and interactions are lost. At a smaller scale, loops are rewired characterised by the gain of longer loops with stronger insulation. Thereby, the results suggest that active RNAPII can restrain loop extrusion. In Chapter 2, in situ Micro-C revealstranscription-based loops, which arise from cis-regulatory contacts at sub-kb resolution. Loss of RNAPII results in weakened loops and affects especially enhancer anchored loops whereas promoter-anchored loops are more stable. I show that RNAPII is required to establish enhancer-anchored loops. Taken together the data reveal the importance of RNAPII to reestablish the chromatin architecture following exit from mitosis, as well as its previously unknown impact on loop extrusion.
Keywords: RNA polymerase II; Cohesin; Chromatin; Epigenetics; 3D chromatin organisation
 
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