Co-Orientation of Replication and Transcription Preserves Genome Integrity

TLDR

Bacterial genomes exhibit a bias toward co‑orientation of replication and transcription, especially for essential or highly expressed genes, a feature that may arise from selection for rapid replication or for preserving genome integrity. The study aimed to distinguish these hypotheses by creating strains with large chromosomal inversions or rRNA‑operon inversions to probe the forces shaping genome organization. New Bacillus subtilis strains were engineered with inversions covering roughly one‑quarter of the chromosome or spanning rRNA operons, enabling comparative analysis of replication dynamics. Mathematical analysis of genomic microarray snapshots revealed that replication is moderately impeded across the inverted region, strongly blocked at inverted rRNA loci in rich medium, triggering DNA damage responses, loss of genome integrity, and cell death, thereby supporting the view that genome‑integrity preservation drives the evolution of replication‑transcription co‑orientation.

Abstract

In many bacteria, there is a genome-wide bias towards co-orientation of replication and transcription, with essential and/or highly-expressed genes further enriched co-directionally. We previously found that reversing this bias in the bacterium Bacillus subtilis slows replication elongation, and we proposed that this effect contributes to the evolutionary pressure selecting the transcription-replication co-orientation bias. This selection might have been based purely on selection for speedy replication; alternatively, the slowed replication might actually represent an average of individual replication-disruption events, each of which is counter-selected independently because genome integrity is selected. To differentiate these possibilities and define the precise forces driving this aspect of genome organization, we generated new strains with inversions either over approximately 1/4 of the chromosome or at ribosomal RNA (rRNA) operons. Applying mathematical analysis to genomic microarray snapshots, we found that replication rates vary dramatically within the inverted genome. Replication is moderately impeded throughout the inverted region, which results in a small but significant competitive disadvantage in minimal medium. Importantly, replication is strongly obstructed at inverted rRNA loci in rich medium. This obstruction results in disruption of DNA replication, activation of DNA damage responses, loss of genome integrity, and cell death. Our results strongly suggest that preservation of genome integrity drives the evolution of co-orientation of replication and transcription, a conserved feature of genome organization.