Polycomb (Pc) group proteins are transcriptional regulators with key roles in development, cell identity and differentiation. Pc-bound chromatin regions form repressive domains that interact in 3D to assemble repressive nuclear compartments. Here, we used multiplexed chromatin imaging to investigate whether Pc compartments involve the clustering of multiple Pc domains during Drosophila development. Notably, 3D proximity between Pc targets is rare and involves predominantly pairwise interactions. These 3D proximities are particularly enhanced in segments where Pc genes are co-repressed. In addition, segment-specific expression of Hox Pc targets leads to their spatial segregation from Pc repressed genes. Finally, non-Hox Pc targets are proximal in regions where they are co-expressed. These results indicate that long-range Pc interactions are temporally and spatially regulated during differentiation and development but do not involve clustering of multiple distant Pc genes.

Controlled motion and positioning of colloids and macromolecular complexes in a fluid, as well as catalytic particles in active environments, are fundamental processes in physics, chemistry and biology. Here we focus on an active biological system for which precise experimental results are available. Our work is fully inspired by studies of one of the most widespread and ancient mechanisms of liquid phase macromolecular segregation and positioning known in nature: bacterial DNA segregation systems. Efficient bacterial chromosome segregation typically requires the coordinated action of a three-component, fueled by adenosine triphosphate machinery called the partition complex. We can distinguish two steps: (i) a process of phase transition [2,3] to built a membraneless region of high protein concentration (partition complex) (ii) the action of molecular motor action upon the complex to create a chemical force.We present a phenomenological model [1] accounting for the dynamics of this system that is also relevant for the physics of catalytic particles in active environments. The model is obtained by coupling simple linear reaction-diffusion equations with a volumetric chemophoresis force field that arises from protein-protein interactions and provides a physically viable mechanism for complex translocation. This description captures experimental observations: dynamic oscillations of complex components, complex separation and symmetrical positioning. The predictions of our model are in agreement with and provide substantial insight into recent experiments. From a non-linear physics view point, this system explores the active separation of matter at micrometric scales with a dynamical instability between static positioning and travelling wave regimes triggered by the dynamical spontaneous breaking of rotational symmetry. We also discuss the phase transition mechanism giving rise to macromolecular assembly of proteins. Our predictions are compared to Super Resolution microscopy and microbiology experiments [1,2,3].[1] Walter J.-C., Dorignac J., Lorman V., Rech J., Bouet J.-Y., Nollmann M., Palmeri J., Parmeggiani A. and Geniet F., Phys. Rev. Lett. 119, 028101 (2017).[2] Debaugny R., Sanchez A., Rech J., Labourdette D., Dorignac J., Geniet F., Palmeri J., Parmeggiani A., Boudsocq, Leberre V., Walter* J.-C. and Bouet* J.-Y Mol. Syst. Biol. 14, e8516 (2018).[3] David G., Walter J.-C., Broedersz C., Dorignac J., Geniet F., Parmeggiani A., Walliser N.-O. and Palmeri J., Phys. Rev. Res. 2, 033377 (2020).

The last decade has seen a spectacular development of genomic physics both in bacteria and eukaryotes. This is due in large part to the development of experimental techniques, which allow to probe (i) the motion of nucleic acids (chromosomal DNA and RNA) in vivo by means of microscopy techniques and (ii) the interactions of these nucleic acids with other actors of the cell (like proteins and molecular motors) by means of high-throughput sequencing techniques. I will focus on three different processes : (i) the modeling of translation with a ballistic model describing the motion of ribosomes on messenger RNA, (ii) the physical mechanism of liquid-liquid phase separation, which is found to be universally spread in the cells to increase, locally and transiently, the concentration of a molecular actor (proteins, RNA etc.). We illustrate this process with the example of the bacterial DNA segregation where liquid-liquid phase separation is used to increase the local concentration in order to catalyze ATP hydrolysis. Finally (iii) I give a general framework on how polymer physics can help to infer the organization of DNA in vivo from microscopy and high-throughput sequencing experiments and apply it to the modeling of bacterial DNA in Escherichia coli and to the epigenetic regulation via polycomb domains in Drosophila.

The bacterial flagellar motor (BFM) is the membrane-embedded rotary molecular motor which turns the flagellum that provides thrust to many bacterial species. This large multimeric complex, composed of a few dozen constituent proteins, has emerged as a hallmark of dynamic subunit exchange. The stator units are inner-membrane ion channels which dynamically bind and unbind to the peptidoglycan at the rotor periphery, consuming the ion motive force (IMF) and applying torque to the rotor when bound. The dynamic exchange is known to be a function of the viscous load on the flagellum, allowing the bacterium to dynamically adapt to its local viscous environment, but the molecular mechanisms of exchange and mechanosensitivity remain to be revealed. Here, by actively perturbing the steady-state stator stoichiometry of individual motors, we reveal a stoichiometry-dependent asymmetry in stator remodeling kinetics. We interrogate the potential effect of next-neighbor interactions and local stator unit depletion and find that neither can explain the observed asymmetry. We then simulate and fit two mechanistically diverse models which recapitulate the asymmetry, finding stator assembly dynamics to be particularly well described by a two-state catch-bond mechanism.

erratum

The ParABS system is composed of three components: a centromeric sequence, parS, and two proteins, the ParA (ATPase) and the ParB DNA binding proteins. Hundreds of ParB proteins assemble dynamically to form nucleoprotein parS-anchored complexes (ParBS) that serve as substrates for ParA molecules to catalyze positioning and segregation events. We address the theoretical modeling of ParBS with a Lattice Gas models and compare it to experiments [1]. We give evidences that (i) ParBS is formed via a Liquid-Liquid Phase Separation in the metastable region of the phase diagram and (ii) this nucleation is catalyzed with the parS sequence [1].[1] Guilhas, Walter,... & Nollmann (2020). ATP-driven separation of liquid phase condensates in bacteria. Mol. Cell, 79, 293-303.

ParAB S , the most widespread bacterial DNA segregation system, is composed of a centromeric sequence, parS , and two proteins, the ParA ATPase and the ParB DNA binding proteins. Hundreds of ParB proteins assemble dynamically to form nucleoprotein parS -anchored complexes that serve as substrates for ParA molecules to catalyze positioning and segregation events. The exact nature of this ParB S complex has remained elusive, what we address here by revisiting the Stochastic Binding model (SBM) introduced to explain the non-specific binding profile of ParB in the vicinity of parS . In the SBM, DNA loops stochastically bring loci inside a sharp cluster of ParB. However, previous SBM versions did not include the negative supercoiling of bacterial DNA, leading to use unphysically small DNA persistences to explain the ParB binding profiles. In addition, recent super-resolution microscopy experiments have revealed a ParB cluster that is significantly smaller than previous estimations and suggest that it results from a liquid-liquid like phase separation. Here, by simulating the folding of long (≥ 30 kb) supercoiled DNA molecules calibrated with realistic DNA parameters and by considering different possibilities for the physics of the ParB cluster assembly, we show that the SBM can quantitatively explain the ChIP-seq ParB binding profiles without any fitting parameter, aside from the supercoiling density of DNA, which, remarkably, is in accord with independent measurements. We also predict that ParB assembly results from a non-equilibrium, stationary balance between an influx of produced proteins and an outflux of excess proteins, i.e., ParB clusters behave like liquid-like protein condensates with unconventional “leaky” boundaries.