IPLS Seminar - Prof. Ethan Garner (Harvard University)

Abstract: How cells specify their dimensions and change their rate of growth in response to nutrient conditions remains an outstanding question. The shape of bacteria is determined by their cell wall, a crosslinked macromolecule holding them in shape. To grow in defined shapes, bacteria must add material into this structure in a spatially controlled manner, and change the rate of insertion to modulate their rate of growth. Two spatially distinct enzymatic systems mediate the growth of the cell wall: The Rod complex, oriented by MreB filaments, moves around the cell circumference, while class A penicillin-binding proteins (aPBPs) do not. To understand how these two systems control bacterial shape and growth, we combine optical assays of enzyme activity with genetic perturbations and computational analysis. 

We find that the diameter of rods is not determined by MreB, rather it depends on the balance between the systems: The Rod complex reduces diameter, while aPBPs increase it. The enzyme pair within the Rod complex (RodA/PBP2A) can both thin or widen cells, depending on its levels relative to MreBCD. Increased Rod complex activity correlates with an increased density of directional MreB filaments and a higher fraction of directionally moving cell wall synthesis enzymes. This increased circumferential synthesis increases the relative amount of oriented material within the sacculi, which increases the structural anisotropy of the sacculi such that they are more resistant to stretching across their width, reinforcing the rod-shape.

Furthermore, we find that cells regulate the density of MreB filaments (but not their speed) in response to nutrient availability. These filaments then control the directional motion and activity of their associated enzymes. Our experiments revealed that the cell regulates the number of MreB filaments by reading out the cell wall precursor lipid II, increasing or decreasing the enzymatic pathway responsible for lipid II synthesis controls both the number of MreB filaments as well as the rate of growth. Lipid II levels are sensed by the serine/threonine kinase PrkC, which phosphorylates a variety of targets, including RodZ, a putative MreB filament nucleator. Surprisingly, we found that strains that overexpress PrkC or contain the phosphomimetic RodZ mutation grow 30%-50% faster than they would normally in the same media. These experiments demonstrate that bacteria regulate their rate of growth by measuring the flux of peptidoglycan precursors through cell wall synthesis.