Final Report Summary - CDNF (Compartmentalization and dynamics of Nuclear functions)
The eukaryotic genome is packaged into chromatin as large structures that occupy distinct domains in the cell nucleus. This organization is thought to be crucial for the normal functions of the genome and, hence, of the cell; defects in architectural elements of the nucleus, for example, are responsible for several human diseases. Not only is the genome organized in three dimensions, but changes occurs in its spatial organization throughout the cell cycle, at different stages of development, and in different metabolic states. Thus, the intimate link between nuclear architecture, nuclear functions and their dynamics, emerges as an important area. However, the causal relationship between nuclear organization and function remains often elusive.
To address this issue, we use budding yeast as a powerful model system that allows to combine effectively genetics, advanced live microscopy and systems biology approaches. Over the last two decades yeast has proven to be an excellent model system for testing the functional role of higher-order chromatin organization (Taddei & Gasser 2012). Using this model system, we ask two main questions:
(1) What determines the spatial and temporal behavior of chromatin
(2) How nuclear organization affects two essential functions of the genome: gene expression and the maintenance of genome integrity.
To address the first question, we focus on the clustering of silent chromatin as a model of functional compartment in which the clustering of repetitive DNAs leads to the sequestration of general repressors of transcription, a phenomenon conserved from yeast to human (Meister & Taddei, 2013). Deciphering how such microenvironments are formed despite the absence of physical barrier to delimitate them, and what regulate their dynamics in relation to changes in genome activity is a key step in understanding how nuclear organization participates in nuclear function. In budding yeast, heterochromatin is mainly found at telomeres that cluster in foci at the nuclear periphery. We have recently shown that the silencing factor Sir3 is a limiting factor for the clustering of telomeres and that Sir3 can promote telomere clustering independently of heterochromatin formation (Ruault et al., 2011). In order to gain insight into the physical mechanisms underlying the dynamics of silent chromatin we integrate our experimental data into quantitative models generating hypothesis that are then tested experimentally (Hoze et al., 2013).
To address the second question, we have developed tools to score and manipulate chromatin position and dynamics of specific loci and test the impact of these two parameters on gene expression and genome stability (Neuman et al, 2012; Loiodice et al, 2014).
Finally, we have identified a new pathway that links replication stress with the formation of heterochromatin (Dubarry et al., 2011). Such a mechanism may contribute to genome integrity by preventing collisions between the replication and transcription machineries. We now aim at deciphering the molecular mechanisms underlying this pathway.
To address this issue, we use budding yeast as a powerful model system that allows to combine effectively genetics, advanced live microscopy and systems biology approaches. Over the last two decades yeast has proven to be an excellent model system for testing the functional role of higher-order chromatin organization (Taddei & Gasser 2012). Using this model system, we ask two main questions:
(1) What determines the spatial and temporal behavior of chromatin
(2) How nuclear organization affects two essential functions of the genome: gene expression and the maintenance of genome integrity.
To address the first question, we focus on the clustering of silent chromatin as a model of functional compartment in which the clustering of repetitive DNAs leads to the sequestration of general repressors of transcription, a phenomenon conserved from yeast to human (Meister & Taddei, 2013). Deciphering how such microenvironments are formed despite the absence of physical barrier to delimitate them, and what regulate their dynamics in relation to changes in genome activity is a key step in understanding how nuclear organization participates in nuclear function. In budding yeast, heterochromatin is mainly found at telomeres that cluster in foci at the nuclear periphery. We have recently shown that the silencing factor Sir3 is a limiting factor for the clustering of telomeres and that Sir3 can promote telomere clustering independently of heterochromatin formation (Ruault et al., 2011). In order to gain insight into the physical mechanisms underlying the dynamics of silent chromatin we integrate our experimental data into quantitative models generating hypothesis that are then tested experimentally (Hoze et al., 2013).
To address the second question, we have developed tools to score and manipulate chromatin position and dynamics of specific loci and test the impact of these two parameters on gene expression and genome stability (Neuman et al, 2012; Loiodice et al, 2014).
Finally, we have identified a new pathway that links replication stress with the formation of heterochromatin (Dubarry et al., 2011). Such a mechanism may contribute to genome integrity by preventing collisions between the replication and transcription machineries. We now aim at deciphering the molecular mechanisms underlying this pathway.