High-throughput imaging and quantitative analysis uncovers the nature of plasmid positioning by ParABS

Abstract

AbstractThe faithful segregation and inheritance of bacterial chromosomes and low-copy number plasmids requires dedicated partitioning systems. The most common of these, ParABS, consists of ParA, a DNA-binding ATPase and ParB a protein that binds to centromeric-like parS sequences on the DNA cargo. The resulting nucleoprotein complexes are believed to move up a self-generated gradient of nucleoid-associated ParA. However, it remains unclear how this leads to the observed cargo positioning and dynamics. In particular, the evaluation of models of plasmid positioning has been hindered by the lack of quantitative measurements. Here, we use high-throughput imaging, analysis and modelling to determine the dynamical nature of these systems. We find that F plasmid is actively brought to specific subcellular home positions within the cell with dynamics akin to an over-damped spring. We identify ParA hopping on the nucleoid as the primary determinant of this geometry-sensing and develop a unified computational model that quantitatively explains the observed behaviour. Furthermore, we confirm our model prediction that cells with the lowest plasmid concentration transition to oscillatory dynamics and find similar behaviour in a distantly-related ParABS system. Our results indicate that ParABS regularly positions plasmids across the nucleoid but operates just below the threshold of an oscillatory instability, minimising ATP consumption. Overall, this work uncovers the dynamical nature of plasmid positioning by ParABS and provides insights relevant for chromosome-based systems.