Cell Division Site Placement and Asymmetric Growth in Mycobacteria

Abstract

Mycobacteria are members of the actinomycetes that grow by tip extension and lack apparent homologues of the known cell division regulators found in other rod-shaped bacteria. Previous work using static microscopy on dividing mycobacteria led to the hypothesis that these cells can grow and divide asymmetrically, and at a wide range of sizes, in contrast to the cell growth and division patterns observed in the model rod-shaped organisms. In this study, we test this hypothesis using live-cell time-lapse imaging of dividing Mycobacterium smegmatis labelled with fluorescent PBP1a, to probe peptidoglycan synthesis and label the cell septum. We demonstrate that the new septum is placed accurately at mid-cell, and that the asymmetric division observed is a result of differential growth from the cell tips, with a more than 2-fold difference in growth rate between fast and slow growing poles. We also show that the division site is not selected at a characteristic cell length, suggesting this is not an important cue during the mycobacterial cell cycle.

Single-cell dynamics of the chromosome replication and cell division cycles in mycobacteria

“…7b). These observations are inconsistent with a 'strict' size-control mechanism that would trigger cell division once a critical cell size is achieved 16 . Nonetheless, our data suggest that some form of size control must be operational in mycobacteria because the interdivision time of single cells (Fig.…”

“…1a and Supplementary Movie 2). Analysis of time-lapse image series confirms that the septum in mycobacteria is not always placed exactly at midcell 12,15,16 . In the majority of cells, septum position is skewed towards the new pole ( Fig.…”

“…2d) underscores the importance of using validated markers, such as Wag31-GFP, to determine the timing of cytokinesis in single-cell experiments. Several recent studies, including this one, have demonstrated that rod-shaped mycobacteria usually divide asymmetrically, giving rise to sibling cells of unequal sizes [15][16][17] . In most cell divisions, displacement of the septum towards the newly formed cell pole causes the old-pole sibling to be larger than the new-pole sibling at birth.…”

“…Although the mechanism responsible for asymmetric division is unclear, we can envisage two alternative scenarios, which are not mutually exclusive. In the first scenario, the nascent septum is initially localized accurately at midcell; however, preferential elongation of the old cell pole during the period from septum positioning to cytokinesis results in unequally sized sibling cells 15,16 . This explanation seems inconsistent with our observation that both cell poles elongate with the same velocity following cell separation (Fig.…”

“…Recently, it was reported that Mycobacterium smegmatis grows in an unusual unipolar manner by preferential extension of the old cell pole, whereas the new cell pole remains quiescent until the next cell division has been completed 12,15,16 . Unipolar growth was hypothesized to give rise to asymmetric cell division, resulting in sibling cells of unequal sizes and growth velocities, with larger old-pole cells growing faster than their smaller newpole siblings 15 .…”