Post-translational modifications of GlmR integrate metabolic and stress signals to maintain cell envelope homeostasis in Bacillus subtilis.

The metabolic networks of most life forms integrate cost-benefit analysis to properly budget carbon and other essential nutrients. Bacillus subtilis is a Gram-positive model bacterium found in diverse ecological niches such as soil, marine environments, and the human gut. As such, B. subtilis cells fine-tune metabolic pathways by monitoring signals indicating the presence of nutrients and stressors. A highly conserved protein, GlmR, is a key player in rationing carbon for the production of cell envelope precursors. This function of GlmR can be attributed to its role in cell shape regulation and antibiotic resistance. Given its central position in carbon utilization, GlmR is under post-translational regulation by phosphorylation and UDP-N-acetylglucosamine (UDP-GlcNAc) binding. GlmR is also linked to cyclic-di-AMP (c-di-AMP), a nucleotide second messenger involved in osmotic and cell wall stress response. In this study, we probed the importance of GlmR in cell morphogenesis, c-di-AMP signaling, and investigated the physiological significance of post-translational regulation. Our results reveal that cells lacking glmR exhibit: (i) increased susceptibility to tunicamycin, a cell envelope targeting antibiotic; (ii) impaired division site positioning; and (iii) reduced intracellular c-di-AMP concentration. Furthermore, we show that the function of GlmR is fine-tuned by UDP-GlcNAc binding, phosphorylation, and acetylation. Additionally, we provide evidence showing that the recently discovered uridyltransferase activity of GlmR is integral for its function. We show that GlmR is a cell width determinant and propose a model suggesting close cooperation with an actin-like protein, MreB. Overall, our studies highlight the importance of the enzymatic function of GlmR and elucidate the mechanism behind the multiple post-translational means to regulate this crucial protein which is at the crux of carbon flux with an important role in maintaining cell envelope integrity.