Mechanistic insights into the spatial organization of RNA polymerase proteins and the chromosome in E. coli cells

Abstract

Along the bacterial chromosome, regions called rrn operons contain genes that are transcribed into ribosomal RNA. These operons are among the most transcriptionally active sites in the genome. It has been observed in E. coli that RNA polymerase (RNAP), while binding to these genetic loci along the chromosome during transcription, forms dense clusters, leading to spatial colocalization of the operons within the cell. Recent experimental evidence suggests that liquid-liquid phase separation contributes to the formation of RNAP clusters, with the antitermination factor NusA playing a key role. We present a simulation model to investigate the mechanisms underlying the formation of these biomolecular condensates. We propose that mutual attraction between NusA proteins, which exhibit a miscibility gap at higher concentrations, drives condensate formation via a polymer-assisted condensation pathway, and we demonstrate how these condensates promote the colocalization of rrn operons. Our results reconcile seemingly disparate experimental observations of chromosomal organization reported in fluorescence-based imaging and Hi-C experiments.