Condensin- and Replication-Mediated Bacterial Chromosome Folding and Origin Condensation Revealed by Hi-C and Super-resolution Imaging

Abstract

Chromosomes of a broad range of species, from bacteria to mammals, are structured by large topological domains whose precise functional roles and regulatory mechanisms remain elusive. Here, we combine super-resolution microscopies and chromosome-capture technologies to unravel the higher-order organization of the Bacillus subtilis chromosome and its dynamic rearrangements during the cell cycle. We decipher the fine 3D architecture of the origin domain, revealing folding motifs regulated by condensin-like complexes. This organization, along with global folding throughout the genome, is present before replication, disrupted by active DNA replication, and re-established thereafter. Single-cell analysis revealed a strict correspondence between sub-cellular localization of origin domains and their condensation state. Our results suggest that the precise 3D folding pattern of the origin domain plays a role in the regulation of replication initiation, chromosome organization, and DNA segregation.

Structures of partition protein ParA with nonspecific DNA and ParB effector reveal molecular insights into principles governing Walker-box DNA segregation

“…The multifaceted DNA-binding surface revealed in the ParA-AMPPNP-DNA structure indicates that ParA would favor interactions with three-dimensional (3D) DNA arrays, which would lead ParA molecules to become embedded within the volume of the nucleoid. This DNA-binding property is consonant with biochemical data showing that ParA-ATP engages in intersegmental transfer between DNA sites (Vecchiarelli et al 2010) as well as more recent superresolution analyses showing that ParA proteins appeared to localize to high-density DNA regions (HDRs) within chromosomes (Marbouty et al 2015;Le Gall et al 2016). The multidimensional DNA-binding feature revealed in the ParA dimer from the ParA-AMPPNP-DNA structure would therefore negate the requirement for membrane-nucleoid confinement, as DNA-embedded ParA would trap its ParB-DNA cargo in the nucleoid cloud, preventing its escape into the cytosol.…”

“…Here we describe the first structures of ParA-AMPPNP-nsDNA and ParA-AMPPNP-ParB complexes. The resultant structural findings provide key insight into these partition steps that, when combined with previous biochemical in vitro reconstitution (Vecchiarelli et al 2013a(Vecchiarelli et al ,b, 2014 and superresolution (Marbouty et al 2015;Le Gall et al 2016) studies, support a diffusion ratchet-like mechanism. Central to this model is our structural finding that, in the presence of DNA, ParA-ATP assumes a dimer conformation favorable for DNA binding, consistent with the predicted ParA-ATP * state (Vecchiarelli et al 2010).…”

“…Recent 3D superresolution studies revealed that the most prominent bacterial HDRs are located at the ends of the segregating nucleoids near the cell poles (Marbouty et al 2015). Thus, these data suggest a molecular segregation mechanism in which ParA and its ParB-DNA cargo would equilibrate to these prominent HDRs either near the surface of the nucleoid or within the nucleoid (HDR2 in Fig.…”

Cohesins and condensins orchestrate the 4D dynamics of yeast chromosomes during the cell cycle

“…However, the initiation and progression of replication, followed by the segregation of the sister chromatids (SCs) into daughter cells, is expected to modify the genome higher‐order organization. Recent studies have unveiled cell‐cycle stage‐specific genome‐wide topological variations in bacteria, yeast, fly, and mammals (Naumova et al , 2013; Guidi et al , 2015; Marbouty et al , 2015; Hug et al , 2017). As expected, in all species the largest reorganization transition is associated with SC condensation, a fundamental process occurring concomitantly to their individualization, and facilitating their proper segregation.…”

Chromosome conformation capture technologies and their impact in understanding genome function

“…Due to its unbiased approach and the decreasing cost of nextgeneration sequencing, Hi-C has been widely used for the analysis of organizational principles of prokaryotic and eukaryotic genomes, of mitotic chromosome structure and for detection of conformational changes in human disease (Dixon et al 2012;Le et al 2013;Marbouty et al 2015;McCord et al 2013;Naumova et al 2013). Hi-C is the method of choice when one is looking for changes at the TAD or supra-TAD level in chromatin organization.…”