Sidewall cell envelope synthesis and remodeling in pole-growing mycobacteria

Abstract Rod-shaped mycobacteria expand from their poles, yet many of the enzymes for cell envelope synthesis localize to both the poles and sidewall. Since envelope synthesis along the non-growing cell periphery could provide a means by which to edit this essential, multi-layered barrier, we tested whether the enzymes are active outside of elongation and division. Metabolic labeling revealed that peptidoglycan and the mycomembrane are assembled in asymmetric polar gradients that continue along the sidewall. All of the peptidoglycan probes tested were sensitive to l,d-transpeptidase activity, including d-alanine-d-alanine derivatives that we show incorporate into cytoplasmic precursors of the biopolymer. However, sidewall labeling by the dipeptide probe persisted when periplasmic remodeling was inhibited, indicating that peptidoglycan metabolism in this region includes precursor synthesis. Our observations suggest a post-expansion role for envelope assembly along the mycobacterial cell periphery and provide a conceptual framework for understanding cell-wide envelope robustness in the face of polar growth.