Relative Distribution of DnaA and DNA in Escherichia coli Cells as a Factor of Their Phenotypic Variability

Phenotypic variability in isogenic bacterial populations is a remarkable feature that helps them cope with external stresses, yet it is incompletely understood. This variability can stem from gene expression noise and/or unequal partitioning of low-copy-number freely diffusing proteins during cell division. Some high-copy-number components are transiently associated with almost immobile large assemblies (hyperstructures) and may be unequally distributed, contributing to bacterial phenotypic variability. We focus on the nucleoid hyperstructure containing numerous DNA-associated proteins, including the replication initiator DnaA. Previously we found an increasing asynchrony in the nucleoid segregation dynamics in growing E. coli cell lineages and suggested that variable replication initiation timing may be the main cause of this phenomenon. Here we support this hypothesis revealing that DnaA/DNA variability represents a key factor leading to the enhanced asynchrony in E. coli. We followed the intra- and intercellular distribution of fluorescently tagged DnaA and histone-like HU chromosomally encoded under their native promoters. The diffusion rate of DnaA is low, corresponding to a diffusion-binding mode of mobility, but still one order faster than that of HU. The intracellular distribution of DnaA concentration is homogeneous in contrast to the significant asymmetry in the distribution of HU to the cell halves, leading to unequal DNA content of nucleoids and DnaA/DNA ratios in future daughter compartments. Accordingly, the intercellular variabilities of HU concentration (CV=26%) and DnaA/DNA ratio (CV=18%) are high. The variable DnaA/DNA may cause a variable replication initiation time (initiation noise). Asynchronous initiation at different replication origins may, in turn, be the mechanism leading to the observed asymmetric intracellular DNA distribution. Our findings indicate that the feature determining the variability of initiation time in E. coli is the DnaA/DNA ratio rather than separately each of them. We provide a likely mechanism for the loss of segregation synchrony phenomenon.