Descrizione del progetto
Regolazione genica e funzione del genoma 3D
L’organizzazione tridimensionale del genoma all’interno del nucleo, definito genoma 3D (3DG, 3D Genome), è essenziale per la regolazione genica. Gli elementi costitutivi del 3DG sono stati identificati, ma il loro contributo alla regolazione genica non è chiaro. Il progetto FuncDis3D, finanziato dall’UE, si propone di sezionare funzionalmente il 3DG per comprenderne gli effetti sull’espressione genica. L’approccio utilizzato è quello di indurre variazioni nel 3DG mediante la drastica riduzione delle proteine e valutare tali variazioni nel tempo. I dati preliminari mostrano che con l’esaurimento dei regolatori di 3DG, le variazioni avvengono in poche ore. I ricercatori ridurranno una gamma di proteine in cellule pluripotenti e differenziate per stabilire il loro contributo all’organizzazione del 3DG. La contemporanea caratterizzazione dell’epigenoma e del trascrittoma permetterà loro di stabilire la sequenza delle variazioni.
Obiettivo
The three-dimensional organization of the genome inside the nucleus (3DG) is crucial for gene regulation. Although the fundamental building blocks of the 3DG have been identified, how they contribute to gene regulation is incompletely understood. In recent years my lab has started to functionally dissect the 3DG in order to understand the effects on gene expression. The current proposal intends to expand this work and aims to i) broaden the array of proteins that are known organize the 3DG, ii) determine how loss of these proteins affects the 3DG and iii) elucidate how this influences gene expression.
The key to this is to induce changes in the 3DG by acute protein depletion and measuring changes over time. We have preliminary data showing that by depleting regulators of the 3DG we can induce changes within a few hours. We will deplete a range of proteins in pluripotent and differentiated cells to determine their contribution to 3DG organization. Simultaneous characterization of the epigenome and transcriptome will allow us to determine the sequence of changes. In addition, we will perform Thousands of Reporters Integrated in Parallel (TRIP) after perturbation of the 3DG. TRIP reporters all have the same minimal promoter that acts as a sensor for promoter-enhancer (mis)-communication. By combining TRIP with 4C we can measure the changes in the reporter-enhancer interaction landscape. Our data will generate fundamental insight into the parameters that govern the interactions between promoters and regulatory elements, within the context of the 3DG, and their effects on gene expression. Furthermore, it allows us to determine the temporal order of changes in the 3DG, the epigenome and gene expression, which is essential for the establishment of cause-and-effect relationships. Collectively, our data will lead to a better appreciation of how distal and proximal regulatory regions cooperate to establish cell-type specific gene expression programs.
Campo scientifico
Programma(i)
Argomento(i)
Meccanismo di finanziamento
ERC-COG - Consolidator GrantIstituzione ospitante
1066 CX Amsterdam
Paesi Bassi