Single Cell and Single-Molecule Assays Reveal Bacteria Regulate their Rate of Growth by Actively Reading Out the Level of Cell Wall Precursors

Abstract

How cells regulate their growth rate in response to nutrient conditions remains an outstanding question in biology. Bacteria are surrounded by a peptidoglycan cell wall that holds their shape. This structure is built by MreB filaments that regulate the enzymes that synthesize the cell wall. Collectively termed the Rod Complex, these machines moving around the cell as they insert new material. To understand how cells grow, we used Structured Illumination Microscopy (SIM) and single-molecule tracking to quantitate the number of Rod complexes in Bacillus subtilis growing in different media. First, we found that cells regulate the filament density but not the speed in response to nutrient availability. The directional motion of enzymes is regulated by MreB filaments and their motions transitioning into a diffusive state when MreB filaments are disrupted. Second, we demonstrated that the number of MreB filaments is regulated by the cell wall precursor lipid II, as increased expression of key Mur enzymes that build lipid II increases the number of MreB filaments and the rate of growth. Furthermore, we showed that lipid II is sensed by the serine/theonine kinase PrkC, which phosphorylates RodZ, a MreB filament nucleator. Surprisingly we found that strains containing a phosphomimetic RodZ mutation or overexpression of PrkC causes cells to grow faster than they would in carbon limited media. Together these demonstrate that bacteria regulate their rate of growth by measuring the flux of peptidoglycan precursors through cell wall synthesis.