Using a single-beam, oscillating optical tweezers, we demonstrate trapping and rotation of rod-shaped bacterial cells with respect to the optical axis. The angle of rotation, θ, is determined by the amplitude of the oscillation. It is shown that θ can be measured from the longitudinal cell intensity profiles in the corresponding phase-contrast images. The technique allows viewing the cell from different perspectives and can provide a useful tool in fluorescence microscopy for the analysis of three-dimensional subcellular structures.

The relaxation of a single DNA molecule is studied. The experimental system consists of optical tweezers and a micron-sized bead that is tethered to the bottom of the sample by a single double-stranded DNA molecule. The bead slows down the DNA relaxation from a strongly stretched configuration such that it is passing through stretched equilibrium states. This allows for a theoretical description of the relaxation trajectory, which is in good agreement with experiment.

The primary protein of the bacterial Z ring guiding cell division, FtsZ, has recently been shown to engage in intriguing self-organization together with one of its natural membrane anchors, FtsA. When co-reconstituted on flat supported membranes, these proteins assemble into dynamic chiral vortices whose diameters resemble the cell circumference. These dynamics are due to treadmilling polar FtsZ filaments, supposedly destabilized by the co-polymerizing membrane adaptor FtsA, thus catalysing their turnover. Here we show that FtsA is in fact dispensable and that the phenomenon is an intrinsic property of FtsZ alone when supplemented with a membrane anchor. The emergence of these chiral dynamic patterns is critically dependent on GTP concentration and FtsZ surface densities, in agreement with theoretical predictions. The interplay of membrane tethering, GTP binding, and hydrolysis promotes both, the assembly and the destabilization of FtsZ polymers, leading to the observed treadmilling dynamics. Notably, the vortex chirality is defined by the position of the membrane targeting sequence (mts) and can be inverted when attaching it to the opposite end of FtsZ. This reveals a so far unknown vectorial character of these cytomotive filaments, comprising three orthogonal directions: Filament polarity, curvature, and membrane attachment.

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 the 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 the unequal DNA content of nucleoids and DnaA/DNA ratios in future daughter compartments. Accordingly, the intercellular variabilities in 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 the initiation time in E. coli is the DnaA/DNA ratio, rather than each of them separately. We provide a likely mechanism for the ‘loss of segregation synchrony’ phenomenon.