DNA Replication Initiation Is Blocked by a Distant Chromosome–Membrane Attachment

• Published in: Current biology Vol. 25; no. 16; pp. 2143 - 2149
• Main Authors: Magnan, David, Joshi, Mohan C., Barker, Anna K., Visser, Bryan J., Bates, David
• Format: Journal Article
• Language: English
• Published: England Elsevier Ltd 17.08.2015
• Subjects: Cell Membrane - metabolism; Chromosomes, Bacterial - metabolism; DNA Replication; DNA, Bacterial - chemistry; DNA, Bacterial - genetics; DNA, Bacterial - metabolism; Escherichia coli - chemistry; Escherichia coli - genetics; Escherichia coli - metabolism; Replication Origin
• ISSN: 0960-9822; 1879-0445
• DOI: 10.1016/j.cub.2015.06.058

Although it has been recognized for several decades that chromosome structure regulates the capacity of replication origins to initiate, very little is known about how or if cells actively regulate structure to direct initiation [1–3]. We report that a localized inducible protein tether between the chromosome and cell membrane in E. coli cells imparts a rapid and complete block to replication initiation. Tethers, composed of a trans-membrane and transcription repressor fusion protein bound to an array of operator sequences, can be placed up to 1 Mb from the origin with no loss of penetrance. Tether-induced initiation blocking has no effect on elongation at pre-existing replication forks and does not cause cell or DNA damage. Whole-genome and site-specific fluorescent DNA labeling in tethered cells indicates that global nucleoid structure and chromosome organization are disrupted. Gene expression patterns, assayed by RNA sequencing, show that tethering induces global supercoiling changes, which are likely incompatible with replication initiation. Parallels between tether-induced initiation blocking and rifampicin treatment and the role of programmed changes in chromosome structure in replication control are discussed. •A chromosome to cell membrane tether immediately blocks replication initiation•Tether locations >1 Mb from the replication origin are lethal•The initiation blocking mechanism only functions in cis•Tethering reduces global chromosome supercoiling and expands the nucleoid Magnan et al. show that an induced protein attachment (tether) between the E. coli chromosome and its cell membrane causes an immediate block to replication initiation. The blocking mechanism involves a global change in chromosome structure, which may reflect initiation regulatory strategies in normal cells.