ChromaSTORM · Visualising how proteins fold DNA into topologically associating domains in single human cells
Horizon 2020 — Marie Skłodowska-Curie Actions
- Duration
- 2020-09-01 → 2022-08-31
- EU contribution
- €174,806
- Participants
- 1
- Scheme
- MSCA-IF
Lines connect the coordinator with its partners.
Results in brief
Visualising how proteins fold DNA into topologically associating domains in single human cells
Our body is composed of a multitude of cells with different functions like blood cells, skin cells and neuronal cells. Yet the genetic information in all our cells is the same. How can different phenotypes emerge from cells carrying the identical genetic sequence? It is one of today’s great challenges to understand how genetic information is modulated inside the cell nucleus. Over the last decade, research has revealed that our genome is organised on multiple levels with the help of structuring proteins, and that this spatial organisation regulates the core functions of the genome resulting in specialised cell types. In this context, topologically associating domains (TADs) were identified as fundamental and functional building blocks of chromatin organisation playing important roles in gene regulation and DNA replication. TADs are self-interacting regions with the size of a few hundreds of kilobases (kb). These domains are organised by the two structuring proteins Cohesin and CTCF, and are supposed to be formed by a mechanism called ‘DNA loop extrusion’. Most of our current insight is based on bulk measurements using genome-wide proximity-based ligation approaches. In order to shed light on the spatial organisation of single TADs together with their structuring proteins in single cells this project set out to establish a novel imaging approach using super-resolution microscopy. Recent advances in this technology enable the visualisation of these fine scale genomic structures and their functional dynamics in 3D in single cells at nanometre resolution.
Data: CORDIS, © European Union
Project objective
How can phenotypic variations emerge from cells that carry the same genetic information? It is one of today’s great challenges to understand how genetic information is modulated inside the cell. Over the last decade, insight from genome-wide proximity-based ligation approaches revealed that the genome is organised in a hierarchical manner with the help of structuring proteins, and that this spatial organisation regulates the core functions of the genome, such as transcription, replication and repair. In this context, topologically associating domains (TADs) were identified as fundamental and functional building blocks of chromatin organisation above the nucleosome level. Despite its vital importance, our current understanding of the spatial organisation of TADs remains largely enigmatic. With the advent of super-resolution microscopy, tools are now available for studying genomic structures and their functional dynamics in situ at a resolution of 10 nm which corresponds to the size of a few nucleosomes.The goal of this project is to reveal the principles of TAD organisation in human cells. To achieve this, I will employ a multidisciplinary imaging-based approach. I will simultaneously visualise the DNA backbone of TADs and architectural proteins involved in TAD structure applying 3D super-resolution microscopy. I will focus on the key TAD organisers CTCF and Cohesin, as well as on Mediator and Condensin II. Additionally, I will directly study their individual structuring function for TADs by their acute depletion. Integrating these data with quantitative measurements of absolute protein copy numbers, I will derive a data-driven model of inner TAD organisation in cells. These studies will provide the first 3D description of this fundamental chromatin super-structure and will further our understanding of genome architecture which is a prerequisite for understanding genome function.
Original text from CORDIS.
Participants
- EUROPEAN MOLECULAR BIOLOGY LABORATORY · HeidelbergCoordinatorGermany
Links
Data: CORDIS, © European Union