Topological domain structure of the Escherichia coli chromosome

Abstract

The circular chromosome of Escherichia coli is organized into independently supercoiled loops, or topological domains. We investigated the organization and size of these domains in vivo and in vitro. Using the expression of >300 supercoiling-sensitive genes to gauge local chromosomal supercoiling, we quantitatively measured the spread of relaxation from double-strand breaks generated in vivo and thereby calculated the distance to the nearest domain boundary. In a complementary approach, we gently isolated chromosomes and examined the lengths of individual supercoiled loops by electron microscopy. The results from these two very different methods agree remarkably well. By comparing our results to Monte Carlo simulations of domain organization models, we conclude that domain barriers are not placed stably at fixed sites on the chromosome but instead are effectively randomly distributed. We find that domains are much smaller than previously reported, ∼10 kb on average. We discuss the implications of these findings and present models for how domain barriers may be generated and displaced during the cell cycle in a stochastic fashion.

The bacterial nucleoid: A highly organized and dynamic structure

“…The length of individual supercoiled domains thereby follows an exponential distribution with a mean value of about 10 kb, which corresponds to approximately 500 domains per chromosome. These computationally derived results are in good agreement with values obtained by analyzing the lengths of individual supercoiled loops on electron micrographs of gently isolated E. coli nucleoids [Postow et al, 2004].…”

“…They rather appear to be placed in a completely random way, so that their positions vary stochastically within a population of cells [Higgins et al, 1996;Deng et al, 2004]. These findings, which were based on probing small, defined regions of the chromosome, were corroborated by a whole genome approach aimed at determining the length and distribution of topological domains in E. coli on a global scale [Postow et al, 2004]. To this end, supercoils were relaxed in live cells using an inducible rare-cutting restriction endonuclease.…”

“…The existence of small and dynamically positioned topological domains provides several advantages for bacteria [Postow et al, 2004]. First, lesions in the chromosome caused by DNA damage, repair processes, or replication only relax one single domain without affecting the superhelicity of other DNA regions.…”

“…Short distances between domain boundaries might, in addition, facilitate the repair of double-strand breaks because they restrict the movement of the two ends to be joined. By concentrating catenane links and thus increasing the free energy of catenanes, they further improve the efficiency of chromosome unlinking at the end of DNA replication [Postow et al, 2004]. Finally, organization into small supercoiled modules significantly reduces the chromosomal radius of gyration, thereby making an important contribution to the overall compaction of bacterial chromatin [Trun and Marko, 1998].…”

Large-Scale Conformational Transitions in Supercoiled DNA Revealed by Coarse-Grained Simulation

“…In Escherichia coli and other bacteria, the circular DNA chromosome is regulated in an underwound topological linking state by topoisomerases, which gives rise to supercoiling of the chromosome. The degree of supercoiling is dynamically adapted to environmental cues (4), and the chromosome is further dynamically partitioned into looped topological domains with average sizes of~10 kilobases (5)(6)(7). In both eukaryotes and prokaryotes, supercoiling facilitates compaction of the genome into the cell (1) and is believed to be intimately connected to regulation of gene expression (7)(8)(9).…”