Posttranslational Control of PlsB Is Sufficient To Coordinate Membrane Synthesis with Growth in Escherichia coli

Abstract

Every cell must produce enough membrane to contain itself. However, the mechanisms by which the rate of membrane synthesis is coupled with the rate of cell growth remain unresolved. By comparing substrate and enzyme concentrations of the fatty acid and phospholipid synthesis pathways of Escherichia coli across a 3-fold range of carbon-limited growth rates, we show that the rate of membrane phospholipid synthesis during steady-state growth is determined principally through allosteric control of a single enzyme, PlsB. Due to feedback regulation of the fatty acid pathway, PlsB activity also indirectly controls synthesis of lipopolysaccharide, a major component of the outer membrane synthesized from a fatty acid synthesis intermediate. Surprisingly, concentrations of the enzyme that catalyzes the committed step of lipopolysaccharide synthesis (LpxC) do not differ across steady-state growth conditions, suggesting that steady-state lipopolysaccharide synthesis is modulated primarily via indirect control by PlsB. In contrast to steady-state regulation, we found that responses to environmental perturbations are triggered directly via changes in acetyl coenzyme A (acetyl-CoA) concentrations, which enable rapid adaptation. Adaptations are further modulated by ppGpp, which regulates PlsB activity during slow growth and growth arrest. The strong reliance of the membrane synthesis pathway upon posttranslational regulation ensures both the reliability and the responsiveness of membrane synthesis. IMPORTANCE How do bacterial cells grow without breaking their membranes? Although the biochemistry of fatty acid and membrane synthesis is well known, how membrane synthesis is balanced with growth and metabolism has remained unclear. This is partly due to the many control points that have been discovered within the membrane synthesis pathways. By precisely establishing the contributions of individual pathway enzymes, our results simplify the model of membrane biogenesis in the model bacterial species Escherichia coli. Specifically, we found that allosteric control of a single enzyme, PlsB, is sufficient to balance growth with membrane synthesis and to ensure that growing E. coli cells produce sufficient membrane. Identifying the signals that activate and deactivate PlsB will resolve the issue of how membrane synthesis is synchronized with growth.

Malonyl-acyl carrier protein decarboxylase activity promotes fatty acid and cell envelope biosynthesis in Proteobacteria

“…The reactions were performed with nonradioactive substrates, the samples were acid precipitated, and the pellets hydrolyzed with Asp-N protease. This process liberates a tripeptide harboring the prosthetic group and attached acyl chains that were then separated and detected by LC-MS/MS ( 17 , 18 , 19 ). Acetyl-ACP was not detected in assays without MadA and was robustly detected in assays containing MadA ( Fig.…”

“…These enzymes produce acetyl-ACP that was first detected as a component of the ACP thioester pool decades ago based on metabolic labeling and electrophoretic analyses in E. coli ( 5 , 9 ). More recent mass spectrometry methods confirm acetyl-ACP is a major component of the E. coli ACP pool ( 18 , 19 ). The origin of the acetyl-ACP pool has been a mystery, and the discovery of MadA identifies an enzyme responsible for acetyl-ACP synthesis in Proteobacteria.…”

Branched-chain amino acid metabolism controls membrane phospholipid structure in Staphylococcus aureus

“…Samples were prepared for acyl-ACP quantification as previously described ( 53 ) with modifications. One milliliter aliquots of cultures at midlog phase growth were removed to a 1.5 ml Eppendorf tube containing 250 μl ice cold 10% trichloroacetic acid.…”

Cell-envelope synthesis is required for surface-to-mass coupling, which determines dry-mass density inBacillus subtilis

“…How membrane synthesis and cell-wall synthesis are linked to biomass growth is a question that remains fundamentally not understood in any bacterium. The first committed steps of fatty-acid and phospholipid synthesis are likely the major pathway elements for the control of membrane synthesis (Cronan Jr and Rock, 2008;Dowhan, 2013;Noga et al, 2020;Rock and Cronan, 1996;Cronan Jr and Waldrop, 2002). However, the signals responsible for controlling their activities largely remain to be identified (Noga et al, 2020).…”

“…The first committed steps of fatty-acid and phospholipid synthesis are likely the major pathway elements for the control of membrane synthesis (Cronan Jr and Rock, 2008;Dowhan, 2013;Noga et al, 2020;Rock and Cronan, 1996;Cronan Jr and Waldrop, 2002). However, the signals responsible for controlling their activities largely remain to be identified (Noga et al, 2020). Furthermore, while multiple gene-regulatory feedbacks for cell-wall metabolism have been identified (Patel et al, 2020) Garner (2020) suggested that PrkC senses the availability of lipid II, the precursor of peptidoglycan synthesis, and thus regulates the number of moving MreB filaments.…”