Regulation of Cell Wall Synthesis and Role of Surface Sensing in Bacillus Subtilis

Abstract

A bacterial cell is a self-replicating machine that must, during steady-state growth, duplicate all its contents within its doubling time. To achieve this, bacterial cells acquire nutrients from the environment to obtain metabolic precursors and energy and then allocate these resources to synthesize different macromolecules, such as proteins, DNA, membranes, and external protective structures. Although the biochemical details of how bacteria make these different macromolecules are known, how they allocate their precious resources and regulate the rate at which different macromolecules are synthesized is poorly understood. We explore how the gram-positive bacterium Bacillus subtilis modulates the rates of cell wall and flagellar synthesis. Most bacteria are surrounded by a peptidoglycan cell wall, a covalently crosslinked meshwork of glycans surrounding cells that confers shape and protection from ¬lysis. Interestingly, we find that we can increase the growth rate of B. subtilis in minimal media by over-activating the cell wall synthesis pathway by overexpressing the serine/threonine kinase PrkC, introducing a phosphomimetic mutation in RodZ (a PrkC substrate), or overexpressing MurAA, which catalyzes the first committed step in cell wall biosynthesis. Despite these perturbations affecting components primarily involved in cell wall synthesis, the increase in growth rate is associated with an increase in other essential processes, namely protein synthesis, the frequency of division, and DNA synthesis. This suggests that some sort of feedback connects the rate of cell wall synthesis with other essential processes in B. subtilis, likely involving PrkC. Next, I discuss flagella and the physiological effects of cell trapping on the hour timescale. When B. subtilis is trapped underneath an agarose pad, it increases its cell length by suppressing division while continuing to grow. This increase in cell length is associated with a decrease in the expression of late-stage motility genes, such as the flagellar filament protein Hag, one of the most abundant proteins in B. subtilis. The decrease in the expression of motility genes is necessary for an increase in homeostatic cell length. Together, these data underscore the extent to which the regulation of large macromolecular structures, namely the cell wall and flagella, can affect cell physiology.