ERC FUNDED PROJECTS

Faithful repair of double stranded DNA breaks (DSBs) is essential, as they are at the origin of genome instability, chromosomal translocations and cancer. Cells repair DSBs through different pathways, which can be faithful or mutagenic, and the balance between them at a given locus must be tightly regulated to preserve genome integrity. Although, much is known about DSB repair factors, how the choice between pathways is controlled within the nuclear environment is not understood. We have shown that nuclear architecture and non-random genome organization determine the frequency of chromosomal translocations and that pathway choice is dictated by the spatial organization of DNA in the nucleus. Nevertheless, what determines which pathway is activated in response to DSBs at specific genomic locations is not understood. Furthermore, the impact of 3D-genome folding on the kinetics and efficiency of DSB repair is completely unknown. Here we aim to understand how nuclear compartmentalization, chromatin structure and genome organization impact on the efficiency of detection, signaling and repair of DSBs. We will unravel what determines the DNA repair specificity within distinct nuclear compartments using protein tethering, promiscuous biotinylation and quantitative proteomics. We will determine how DNA repair is orchestrated at different heterochromatin structures using a CRISPR/Cas9-based system that allows, for the first time robust induction of DSBs at specific heterochromatin compartments. Finally, we will investigate the role of 3D-genome folding in the kinetics of DNA repair and pathway choice using single nucleotide resolution DSB-mapping coupled to 3D-topological maps. This proposal has significant implications for understanding the mechanisms controlling DNA repair within the nuclear environment and will reveal the regions of the genome that are susceptible to genomic instability and help us understand why certain mutations and translocations are recurrent in cancer

1 999 750 €

Start date: 2017-03-01, End date: 2022-02-28

This project investigates the two-way relationship between spatio-temporal genome organization and coordinated gene regulation, through an approach at the interface between physics, computer science and biology. In the nucleus, preferred positions are observed from chromosomes to single genes, in relation to normal and pathological cellular states. Evidence indicates a complex spatio-temporal coupling between co-regulated genes: e.g. certain genes cluster spatially when responding to similar factors and transcriptional noise patterns suggest domain-wide mechanisms. Yet, no individual experiment allows probing transcriptional coordination in 4 dimensions (FISH, live locus tracking, Hi-C...). Interpreting such data also critically requires theory (stochastic processes, statistical physics…). A lack of appropriate experimental/analytical approaches is impairing our understanding of the 4D genome. Our proposal combines cutting-edge single-molecule imaging, signal-theory data analysis and physical modeling to study how genes coordinate in space and time in a single nucleus. Our objectives are to understand (a) competition/recycling of shared resources between genes within subnuclear compartments, (b) how enhancers communicate with genes domain-wide, and (c) the role of local conformational dynamics and supercoiling in gene co-regulation. Our organizing hypothesis is that, by acting on their microenvironment, genes shape their co-expression with other genes. Building upon my expertise, we will use dual-color MS2/PP7 RNA labeling to visualize for the first time transcription and motion of pairs of hormone-responsive genes in real time. With our innovative signal analysis tools, we will extract spatio-temporal signatures of underlying processes, which we will investigate with stochastic modeling and validate through experimental perturbations. We expect to uncover how the functional organization of the linear genome relates to its physical properties and dynamics in 4D.

1 499 750 €

Start date: 2018-04-01, End date: 2023-03-31

Although we have extensive knowledge of many important processes in cell biology, including information on many of the molecules involved and the physical interactions among them, we still do not understand most of the dynamical features that are the essence of living systems. This is particularly true for the actin cytoskeleton, a major component of the internal architecture of eukaryotic cells. In living cells, actin networks constantly assemble and disassemble filaments while maintaining an apparent stable structure, suggesting a perfect balance between the two processes. Such behaviors are called “dynamic steady states”. They confer upon actin networks a high degree of plasticity allowing them to adapt in response to external changes and enable cells to adjust to their environments. Despite their fundamental importance in the regulation of cell physiology, the basic mechanisms that control the coordinated dynamics of co-existing actin networks are poorly understood. In the AAA project, first, we will characterize the parameters that allow the coupling among co-existing actin networks at steady state. In vitro reconstituted systems will be used to control the actin nucleation patterns, the closed volume of the reaction chamber and the physical interaction of the networks. We hope to unravel the mechanism allowing the global coherence of a dynamic actin cytoskeleton. Second, we will use our unique capacity to perform dynamic micropatterning, to add or remove actin nucleation sites in real time, in order to investigate the ability of dynamic networks to adapt to changes and the role of coupled network dynamics in this emergent property. In this part, in vitro experiments will be complemented by the analysis of actin network remodeling in living cells. In the end, our project will provide a comprehensive understanding of how the adaptive response of the cytoskeleton derives from the complex interplay between its biochemical, structural and mechanical properties.

2 349 898 €

Start date: 2017-09-01, End date: 2022-08-31

"How are actions initiated by the human brain when there is no external sensory cue or other immediate imperative? How do subtle ongoing interactions within the brain and between the brain, body, and sensory context influence the spontaneous initiation of action? How should we approach the problem of trying to identify the neural events that cause spontaneous voluntary action? Much is understood about how the brain decides between competing alternatives, leading to different behavioral responses. But far less is known about how the brain decides "when" to perform an action, or "whether" to perform an action in the first place, especially in a context where there is no sensory cue to act such as during foraging. This project seeks to open a new chapter in the study of spontaneous voluntary action building on a novel hypothesis recently introduced by the applicant (Schurger et al, PNAS 2012) concerning the role of ongoing neural activity in action initiation. We introduce brain-behavior forecasting, the converse of movement-locked averaging, as an approach to identifying the neurodynamic states that commit the motor system to performing an action "now", and will apply it in the context of information foraging. Spontaneous action remains a profound mystery in the brain basis of behavior, in humans and other animals, and is also central to the problem of asynchronous intention-detection in brain-computer interfaces (BCIs). A BCI must not only interpret what the user intends, but also must detect "when" the user intends to act, and not respond otherwise. This remains the biggest challenge in the development of high-performance BCIs, whether invasive or non-invasive. This project will take a systematic and collaborative approach to the study of spontaneous self-initiated action, incorporating computational modeling, neuroimaging, and machine learning techniques towards a deeper understanding of voluntary behavior and the robust asynchronous detection of decisions-to-act."

1 338 130 €

Start date: 2015-10-01, End date: 2020-09-30