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FROM SINGLE MOLECULES TO CELL REPROGRAMMING: DECIPHERING AND RECODING DISORDERED PIONEER TRANSCRIPTION FACTORS

Project description

Investigation of structure and function of pioneer transcription factors

Pioneer transcription factors (pTFs) initiate cell-fate changes by invading and opening closed chromatin. These unique abilities have been used to regulate cell development and induce pluripotent stem cells. The pTFs consist of structured DNA-binding domains (DBDs) flanked by intrinsically disordered regions (IDRs). Single-molecule spectroscopy allows the monitoring of dynamic molecular systems and measuring their conformational distributions. The ERC-funded PIONEER project aims to uncover the mechanisms of pTFs interactions with nucleosomes, chromatin invading and opening, and the interplay between DBDs and IDRs. Using the known and newly designed pTFs, the study will map conformational states, investigate kinetic mechanisms, and observe chromatin remodelling in vitro and in vivo in the context of four distinct cell reprogramming pathways.

Objective

Pioneer transcription factors (pTFs) have unique capabilities beyond classical TFs: They can invade and open closed chromatin, initiating cell-fate changes. Their remarkable abilities have been used to steer cell-fate decisions and to induce a pluripotent stem cell state through poorly understood pathways. Like most TFs, pTFs consist of structured DNA-binding domains (DBDs) flanked by long intrinsically disordered regions (IDRs). In attempts to explain their pioneering functions, intense focus has been on how the structured DBDs of pTFs interact with the nucleosome core particle. Yet, the critical interactions with nucleosomes beyond the core particle, the interplay between DBDs and IDRs, and the molecular mechanism of chromatin invading and opening, remain unclear.

The extensive disorder of pTFs places them outside the scope of current structural biology efforts and understanding their functions therefore requires a different approach. Single-molecule spectroscopy offers a powerful toolbox to monitor dynamic molecular systems and measure their conformational distributions. These methods enable quantitative modeling of distances and dynamics in biomolecules over timescales reaching over 15 orders of magnitude. Building on our recent breakthroughs in single-molecule techniques for studying highly disordered proteins in chromatin regulation and our preliminary data on pTF IDRs, we are in a unique position to apply our expertise to the molecular mechanism of pTFs.

Using five established pTFs involved in four distinct cell reprogramming pathways, we intend to: 1) map conformational states, 2) decipher kinetic mechanisms, 3) engineer new pTFs, and 4) observe chromatin remodelling, both in vitro and within the complex cellular environment. A molecular-level understanding of pTF functions may break the barrier to fully controlling cell fate, unleashing the enormous medical potential of cell-based therapy.

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(opens in new window) ERC-2021-STG

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Host institution

KOBENHAVNS UNIVERSITET
Net EU contribution

Net EU financial contribution. The sum of money that the participant receives, deducted by the EU contribution to its linked third party. It considers the distribution of the EU financial contribution between direct beneficiaries of the project and other types of participants, like third-party participants.

€ 1 052 761,46
Address
NORREGADE 10
1165 KOBENHAVN
Denmark

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Region
Danmark Hovedstaden Byen København
Activity type
Higher or Secondary Education Establishments
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Total cost

The total costs incurred by this organisation to participate in the project, including direct and indirect costs. This amount is a subset of the overall project budget.

€ 1 052 761,46

Beneficiaries (2)

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