A dynamically assembled cell wall synthesis machinery buffers cell growth

Abstract

Assembly of protein complexes is a key mechanism for achieving spatial and temporal coordination in processes involving many enzymes. Growth of rod-shaped bacteria is a well-studied example requiring such coordination; expansion of the cell wall is thought to involve coordination of the activity of synthetic enzymes with the cytoskeleton via a stable complex. Here, we use singlemolecule tracking to demonstrate that the bacterial actin homolog MreB and the essential cell wall enzyme PBP2 move on timescales orders of magnitude apart, with drastically different characteristic motions. Our observations suggest that PBP2 interacts with the rest of the synthesis machinery through a dynamic cycle of transient association. Consistent with this model, growth is robust to large fluctuations in PBP2 abundance. In contrast to stable complex formation, dynamic association of PBP2 is less dependent on the function of other components of the synthesis machinery, and buffers spatially distributed growth against fluctuations in pathway component concentrations and the presence of defective components. Dynamic association could generally represent an efficient strategy for spatiotemporal coordination of protein activities, especially when excess concentrations of system components are inhibitory to the overall process or deleterious to the cell.

Coarse-grained simulations of bacterial cell wall growth reveal that local coordination alone can be sufficient to maintain rod shape

“…Consistent with our simulations, fluorescently tagged monofunctional transpeptidase PBP2a of Bacillus subtilis moves circumferentially around cells (28,29), and PBP2 of E. coli was shown by bacterial two-hybrid and chemical crosslinking assays to interact with PBP1A in vitro and in the cell (52). A recent study reported, however, that fluorescently-tagged PBP2 of E. coli moves with a diffusive behavior, which is not affected by A22 treatment (70), while the same treatment causes fluorescently-tagged Caulobacter crescentus PBP2 to lose its spiral localization pattern (71). Thus, additional study is needed to resolve these discrepancies.…”

RodZ links MreB to cell wall synthesis to mediate MreB rotation and robust morphogenesis

“…In Escherichia coli, multiple proteins have been suggested to interact with MreB, including the penicillin binding protein (PBP) cell wall synthesis enzymes and RodZ, an integral membrane protein that directly binds MreB (8)(9)(10)(11). PBP2 inhibitors block MreB rotation (5) but the PBP2 protein does not have similar dynamics to MreB (12), thus ruling out PBP2 as the major linker between MreB rotation and cell wall synthesis. In contrast, we discovered that RodZ displays similar dynamics to MreB.…”

Getting into shape: How do rod-like bacteria control their geometry?

“…In fact, some of the cellwall-synthesis enzymes have been seen to move in a similar manner as MreB filaments in the Gram-positive B. subtilis (Garner et al 2011;Domínguez-Escobar et al 2011), supporting the hypothesis of physical interaction. In Gram-negative E. coli, at least one important synthesising enzyme, the transpeptidase PBP2, moves rapidly and diffusively, showing no processivity on the sub-second time scale (Lee et al 2014), thus suggesting a more transient interaction of the cell-wall synthesis proteins. Tuson et al (2012) finds that the timescales at which disrupting MreB affects cell wall elasticity are similar to the growth time, in consistence with this interpretation.…”