Amélie Leforestier

Nanostructures of several ionomers in dispersion in solvents were studied by Small-Angle X-Ray Scattering (SAXS) and cryo-TEM. No size effects were observed between the considered water/alcohol systems, as ionomers assemble into cylindrical primary aggregates of 2 to 3 nm in diameter and a few tenths of nanometers long. Ethylene glycol addition, an additive useful for ink processing, does not impact primary nano-aggregates shape and size but forms agglomerates. The size of these agglomerates is believed to strongly depend on solvent composition and ionomer nature. Schematic representations were proposed from the information gathered from the different methods.

Chromatin organization, through the assembly of DNA with histones and the folding of nucleosome chains, regulates DNA accessibility for transcription, DNA replication and repair. Although models derived from in vitro studies have proposed distinct nucleosome chain geometries, the organization of chromatin within the crowded cell nucleus remains elusive. Using cryo-electron tomography of thin vitreous sections, we directly observed the path of nucleosomal and linker DNA in situ from a flash-frozen organism - Drosophila embryos. We quantified linker length and curvature, characterizing an irregular zig-zag chromatin-folding motif, with a low degree of linker bending. Nucleosome conformations could be identified on individual particles in favorable orientations without structure averaging. Additionally, we observed particles that accommodate a number of DNA gyres ranging from less than one to up to three, which resemble previously proposed non-octameric nucleosomal particles with variable DNA wrapping.

DNA–DNA interactions at work in chromosome shaping and sequence recognition take place in dense and confined environments. Theory and simulation have predicted that, at small separation, interactions between double helices are helically modulated, leading to correlations, a phenomenon poorly documented experimentally. In particular, helical correlations are not compatible with curvature, a general feature of DNA dense states in vitro and in vivo. Using cryo electron microscopy of DNA toroids obtained in vitro in the presence of a condensing agent, we analyze helical correlations in curved condensates. We show that in-phase helix alignments are preferred over a wide range of ionic concentrations. Optimization of helical correlations in a curved assembly is achieved through rearrangements within the toroid as it grows: the nucleation of radial alignments is followed by a circumferential propagation leading to a polygonal shaping of the object. Extended phased flat domains alternate with localized, highly curved, frustrated regions. In addition, the combination of bending and helical correlations results in a local decrease in the DNA helical pitch in highest curvature regions. Overall, this work shows how helical correlations tune both the shape of DNA condensates at the mesoscale and the conformation of the double helix at the nanoscale.

Cellular cryo electron tomography explores tissue and cells in their unstained flash-frozen native state, revealing in situ the structure of macromolecules together with their local environment and interactions with partners, also known as molecular sociology. To obtain thin samples, cryo-FIB milling is nowadays the most popular method, with impressive successes. The alternative, cryo-ultramicrotomy, is often overlooked on account of poorly reproducible attachment of cryo-sections to their support, resulting in extremely low throughput. We optimized the workflow, focusing on section adhesion and their support grids. We thus increased vitreous sections’ cryo electron tomography throughput to equal that of thin film, with typically several tens to hundreds of cryo-tomograms per sample. This open the way to new advances in cellular cryo electron tomography, as the method is devoid of beam damage, can provide large surfaces and serial sections of any type of sample from cells to tissues. In addition, section thickness can be tuned down to 30-50 nm, which may be an advantage for imaging small molecular complexes, such as DNA and nucleosomes.

Imogolite nanotubes present a unique spatial separation of conduction and valence bands but the large bandgap inhibits potential application as a photocatalyst. The first step toward using these well-defined nanotubes in photocatalysis and exploiting their natural polarization-promoting charge separation across the nanotubes wall, is to tune their bandgap energy. Here, titanium modified double walled aluminogermanate imogolite nanotubes are prepared to overcome this limitation. Structural and optical properties are determined at different scales and the photocatalytic performance is evaluated for H2 production under UV-visible light irradiation. Although the incorporation of Ti atoms into the structure remains limited, the resulting nanotubes reveal a remarkable hydrogen evolution rate of ~1500 µmol g-1 h-1, corresponding to a 65-fold increase relative to a reference TiO2P25 photocatalyst. These results confirm the theoretical predictions regarding the untapped potential of modified imogolite nanotubes as promising 1D photoactive polarized nanoreactors.

The chromatin machinery, with DNA-histone assembly and further folding of nucleosome chains, influences DNA availability for functional interactions necessary to the regulation of transcription, DNA replication and repair. Despite models based on in vitro studies, the nucleosome chain geometry within the crowded cell nucleus remains elusive. Using cryo-electron tomography of thin vitreous sections, we directly observed the path of nucleosomal and linker DNA in situ in intact flash-frozen organism - Drosophila embryo. We quantified linker length and curvature characterizing an irregular zig-zag chromatin folding motif, with a low degree of linker bending. Additionally, nucleosome conformational variability with non-canonical structures and non-octameric particles were observed as individual objects, without structure averaging, highlighting the high structural heterogeneity of native chromatin.

In viruses and cells, DNA is closely packed and tightly curved thanks to polyvalent cations inducing an effective attraction between its negatively charged filaments. Our understanding of this effective attraction remains very incomplete, partly because experimental data is limited to bulk measurements on large samples of mostly uncurved DNA helices. Here we use cryo electron microscopy to shed light on the interaction between highly curved helices. We find that the spacing between DNA helices in spermine-induced DNA toroidal condensates depends on their location within the torus, consistent with a mathematical model based on the competition between electrostatic interactions and the bending rigidity of DNA. We use our model to infer the characteristics of the interaction potential, and find that its equilibrium spacing strongly depends on the curvature of the filaments. In addition, the interaction is much softer than previously reported in bulk samples using different salt conditions. Beyond viruses and cells, our characterization of the interactions governing DNA-based dense structures could help develop robust designs in DNA nanotechnologies.

In viruses and cells, DNA is closely packed and tightly curved thanks to polyvalent cations inducing an effective attraction between its negatively charged filaments. Our understanding of this effective attraction remains very incomplete, partly because experimental data is limited to bulk measurements on large samples of mostly uncurved DNA helices. Here we use cryo electron microscopy to shed light on the interaction between highly curved helices. We find that the spacing between DNA helices in spermine-induced DNA toroidal condensates depends on their location within the torus, consistent with a mathematical model based on the competition between electrostatic interactions and the bending rigidity of DNA. We use our model to infer the characteristics of the interaction potential, and find that its equilibrium spacing strongly depends on the curvature of the filaments. In addition, the interaction is much softer than previously reported in bulk samples using different salt conditions. Beyond viruses and cells, our characterization of the interactions governing DNA-based dense structures could help develop robust designs in DNA nanotechnologies.