• Expert ANR

• Membre d'un pool d'experts
• Direction d'équipe

• Physique/Physique/Biophysique
  
• Physique/Matière Condensée/Matière Molle
  
• Physique/Matière Condensée/Mécanique statistique
  

ParABS, 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 ParBS 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.

In response to the fresh water scarcity, Tunisia is utilizing more and more membrane desalination of unconventional resources, including brackish waters and seawater. The widespread reserves of groundwaters and their low salinity make this resource of special interest. Two predominant ionic compositions have been identified depending on their relative proportion of sulfate to chloride ions. The question arising for the decision-makers concerns the choice of membrane technology and, therefore, of membrane. Two nanofiltration (NF) membranes (NF270 and NF90) and a reverse osmosis (RO) one (BW30) were tested in a desalination study of synthetic feeds reproducing the ionic composition of three representative groundwaters. Sulfate/chloride ratio appears to be the key factor for the membrane choice to obtain good quality drinking water meeting the Tunisian standards. Moreover, validation of two prediction tools was investigated: ROSA, software provided by the membrane manufacturer and Nanoflux ® , software specifically designed for NF. The experimental NF results are well fitted by the Nanoflux ® simulations. We concluded that ROSA cannot generally provide good NF predictions because it does not take into account the electric interactions between membrane and feed.

Bacterial ParB partitioning proteins involved in chromosomes and low-copy-number plasmid segregation are CTP-dependent molecular switches. CTP-binding converts ParB dimers to DNA clamps, allowing unidimensional diffusion along the DNA. This sliding property has been proposed to explain the ParB spreading over large distances from parS centromere sites where ParB is specifically loaded. We modeled such a ‘Clamping & sliding’ mechanism as a typical reaction-diffusion system, compared it to the F-plasmid ParB DNA binding pattern, and found that it can account neither for the long range of ParB binding to DNA, nor for the rapid assembly kinetics observed in vivo after parS duplication. Also, it predicts a strong effect on the F-plasmid ParB binding pattern from the presence of a roadblock that is not observed in ChIP-seq. We conclude that although ‘Clamping & sliding’ can occur at short distances from parS, another mechanism must apply for ParB recruitment at larger genomic distances.

We investigate the measurement of DNA supercoiling density ($\sigma$) along chromosomes using interaction frequencies between DNA and DNA-anchored clusters of proteins. Specifically, we show how the physics of DNA supercoiling leads, in bacteria, to the quantitative modeling of binding properties of ParB proteins around their centromere-like site, {\it parS}. Using this framework, we provide an upper bound for $\sigma$ in the {\it Escherichia coli} chromosome, consistent with plasmid values, and offer a proof of concept for a high accuracy measurement. To reach these conclusions, we revisit the problem of the formation of ParB clusters. We predict, in particular, that they result from a non-equilibrium, stationary balance between an influx of produced proteins and an outflux of excess proteins, i.e., they behave like liquid-like protein condensates with unconventional ``leaky'' boundaries.

The cellular cytoplasm is organized into compartments. Phase separation is a simple manner to create membraneless compartments in order to confine and localize particles like proteins. In many cases, these particles are bound to fluctuating polymers like DNA or RNA. We propose a general theoretical framework for such polymer-bound particles and derive an effective 1D lattice gas model with both nearest-neighbor and emergent long-range interactions arising from looped configurations of the fluctuating polymer. We argue that 1D phase transitions exist in such systems for both Gaussian and self-avoiding polymers and, using a variational method that goes beyond mean-field theory, we obtain the complete mean occupation-temperature phase diagram. To illustrate this model, we apply it to the biologically relevant case of ParABS, a prevalent bacterial DNA segregation system.