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High-resolution mapping of the spatial organization of Caulobacter crescentus chromosome by chromosome conformation capture in conjunction with next-generation sequencing (Hi-C)

ABSTRACT: Chromosomes must be highly compacted and organized within cells, but how this is achieved in vivo remains poorly understood. We report the first use of Hi-C to map the structure of bacterial chromosomes at a high, unprecedented resolution. Analysis of Hi-C data and polymer modeling indicates that the Caulobacter crescentus chromosome consists of multiple, largely independent spatial domains likely comprised of supercoiled plectonemes arrayed into a bottlebrush-like fiber. These domains are stable throughout the cell cycle and re-established concomitantly with DNA replication. We provide strong evidence that domain boundaries are established by highly-expressed genes and the formation of plectoneme-free regions. Additionally, we use Hi-C to demonstrate that supercoiling, the histone-like protein HU, and SMC act at different length-scales and in complementary ways to organize and structure chromosomes within cells.

ORGANISM(S): Caulobacter vibrioides CB15

PROVIDER: GSE45966 | GEO | 2013/10/30

SECONDARY ACCESSION(S): PRJNA196826

REPOSITORIES: GEO

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High-resolution mapping of the spatial organization of Caulobacter crescentus chromosome by chromosome conformation capture in conjunction with next-generation sequencing (Hi-C)

Project description:Chromosomes must be highly compacted and organized within cells, but how this is achieved in vivo remains poorly understood. We report the first use of Hi-C to map the structure of bacterial chromosomes at a high, unprecedented resolution. Analysis of Hi-C data and polymer modeling indicates that the Caulobacter crescentus chromosome consists of multiple, largely independent spatial domains likely comprised of supercoiled plectonemes arrayed into a bottlebrush-like fiber. These domains are stable throughout the cell cycle and re-established concomitantly with DNA replication. We provide strong evidence that domain boundaries are established by highly-expressed genes and the formation of plectoneme-free regions. Additionally, we use Hi-C to demonstrate that supercoiling, the histone-like protein HU, and SMC act at different length-scales and in complementary ways to organize and structure chromosomes within cells. Hi-C experiments were performed on untreated wild type swarmer cells and drug-treated swarmer cells (Novobiocin or Rifampicin) of Caulobacter crescentus CB15N. Hi-C were also performed on swarmer cells of smc knock-out and hup1hup2 knock-out mutants of Caulobacter crescentus. Time-course Hi-C were performed on a synchronous population of Caulobacter crescentus at 5, 10, 30, 45, 60 and 75 minutes after synchronization. The progression of DNA replication during the cell cycle was followed by total genomic DNA sequencing.

2013-10-30 | E-GEOD-45966 | biostudies-arrayexpress

Single cell Hi-C reveals cell-to-cell variability in chromosome structure

Project description:Large-scale chromosome structure and spatial nuclear arrangement have been linked to control of gene expression and DNA replication and repair. Genomic techniques based on chromosome conformation capture assess contacts for millions of loci simultaneously, but do so by averaging chromosome conformations from millions of nuclei. Here we introduce single cell Hi-C, combined with genome-wide statistical analysis and structural modeling of single copy X chromosomes, to show that individual chromosomes maintain domain organisation at the megabase scale, but show variable cell-to-cell chromosome territory structures at larger scales. Despite this structural stochasticity, localisation of active gene domains to boundaries of territories is a hallmark of chromosomal conformation, affecting most domains in most nuclei. Single cell Hi-C data bridge current gaps between genomics and microscopy studies of chromosomes, demonstrating how modular organisation underlies dynamic chromosome structure, and how this structure is probabilistically linked with genome activity patterns. Mouse Th1 single-cell Hi-C maps were produced and paired-end sequenced. 10 single-cell samples and a multi-sample pool together with a population Hi-C sample are included.

2013-09-26 | E-GEOD-48262 | biostudies-arrayexpress

Differential roles of positive and negative supercoiling in organizing the E. coli genome

Project description:This study aims to explore whether and how positive and negative supercoiling contribute to the three-dimensional (3D) organization of the bacterial genome. We used recently published Escherichia coli GapR ChIP-seq and TopoI ChIP-seq (also called EcTopoI-seq) data, which marks positive and negative supercoiling sites, respectively, to study how supercoiling correlates with the spatial contact maps obtained from chromosome conformation capture sequencing (Hi-C and 5C). We find that supercoiled chromosomal loci have overall higher Hi-C contact frequencies than sites that are not supercoiled. Surprisingly, positive supercoiling corresponds to higher spatial contact than negative supercoiling. Additionally, positive, but not negative, supercoiling could be identified from Hi-C data with high accuracy. We further find that the majority of positive and negative supercoils coincide with highly active transcription units, with a minor group associated with replication and other genomic processes. Our results show that both positive and negative supercoiling enhance spatial contact, with positive supercoiling playing a larger role in bringing genomic loci closer in space. Based on our results, we propose new physical models of how the E. coli chromosome is organized by positive and negative supercoils.

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Loop-extruding Smc5/6 organizes transcription-induced positive DNA supercoils [RNA-seq]

Project description:The Structural Maintenance of Chromosome (SMC) protein complexes cohesin, condensin and the Smc5/6 complex (Smc5/6) are essential for chromosome function. At the molecular level, these complexes fold DNA by loop extrusion. Accordingly, cohesin creates chromosome loops in interphase, and condensin compacts mitotic chromosomes. However, the role of Smc5/6’s recently discovered DNA loop extrusion activity is unknown. Here, we uncover that Smc5/6 controls the spatial organization of supercoiled chromosomal regions. The results show that Smc5/6 associates with transcription-induced positively supercoiled chromosomal DNA at cohesin-dependent chromosome loop boundaries. Mechanistically, single-molecule imaging reveals that dimers of Smc5/6 specifically recognize the tip of positively supercoiled DNA plectonemes, and efficiently initiates loop extrusion to gather the supercoiled DNA into a large plectonemic loop. Finally, Hi-C analysis shows that Smc5/6 links chromosomal regions containing transcription-induced positive supercoiling in cis. Altogether, our findings indicate that Smc5/6 controls the three-dimensional organization of chromosomes by recognizing and initiating loop extrusion on positively supercoiled DNA.

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Loop-extruding Smc5/6 organizes transcription-induced positive DNA supercoils [Hi-C]

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Loop-extruding Smc5/6 organizes transcription-induced positive DNA supercoils [ChIP-seq]

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Transcription forms and remodels supercoiling domains unfolding large scale chromatin structures [NimbleGen ChIP-chip]

Project description:This study was designed to investigate DNA supercoiling across the human genome and to understand how supercoiling domains impact on higher levels of genome organisation. DNA supercoiling is an inherent consequence of twisting DNA and is critical for regulating gene expression and DNA replication. However, DNA supercoiling at a genomic scale in human cells is uncharacterized. To map supercoiling we used biotinylated-trimethylpsoralen as a DNA structure probe to show the genome is organized into supercoiling domains. Domains are formed and remodeled by RNA polymerase and topoisomerase activities and are flanked by GC-AT boundaries and CTCF binding sites. Under-wound domains are transcriptionally active, enriched in topoisomerase I, M-bM-^@M-^\openM-bM-^@M-^] chromatin fibers and DNaseI sites, but are depleted of topoisomerase II. Furthermore DNA supercoiling impacts on additional levels of chromatin compaction as under-wound domains are cytologically decondensed, topologically constrained, and decompacted by transcription of short RNAs. We suggest that supercoiling domains create a topological environment that facilitates gene activation providing an evolutionary purpose for clustering genes along chromosomes. The binding of bTMP, as a reporter for DNA supercoiling, was investigated in RPE1 cells. Experiments were biological replicates

2013-02-17 | E-GEOD-43448 | biostudies-arrayexpress

Transcription forms and remodels supercoiling domains unfolding large scale chromatin structures [Agilent ChIP-chip]

2013-02-17 | E-GEOD-43450 | biostudies-arrayexpress

Single cell Hi-C reveals cell-to-cell variability in chromosome structure

2013-09-26 | GSE48262 | GEO

Transcription forms and remodels supercoiling domains unfolding large scale chromatin structures [Illumina transcription]

Project description:This study was designed to investigate DNA supercoiling across the human genome and to understand how supercoiling domains impact on higher levels of genome organisation. DNA supercoiling is an inherent consequence of twisting DNA and is critical for regulating gene expression and DNA replication. However, DNA supercoiling at a genomic scale in human cells is uncharacterized. To map supercoiling we used biotinylated-trimethylpsoralen as a DNA structure probe to show the genome is organized into supercoiling domains. Domains are formed and remodeled by RNA polymerase and topoisomerase activities and are flanked by GC-AT boundaries and CTCF binding sites. Under-wound domains are transcriptionally active, enriched in topoisomerase I, M-bM-^@M-^\openM-bM-^@M-^] chromatin fibers and DNaseI sites, but are depleted of topoisomerase II. Furthermore DNA supercoiling impacts on additional levels of chromatin compaction as under-wound domains are cytologically decondensed, topologically constrained, and decompacted by transcription of short RNAs. We suggest that supercoiling domains create a topological environment that facilitates gene activation providing an evolutionary purpose for clustering genes along chromosomes. The gene transcription of 3 independent biological replicates were investigated

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