Peptidoglycan Remodeling Enables Escherichia coli To Survive Severe Outer Membrane Assembly Defect

Abstract

Gram-negative bacteria have a tripartite cell envelope with the cytoplasmic membrane (CM), a stress-bearing peptidoglycan (PG) layer, and the asymmetric outer membrane (OM) containing lipopolysaccharide (LPS) in the outer leaflet. Cells must tightly coordinate the growth of their complex envelope to maintain cellular integrity and OM permeability barrier function. The biogenesis of PG and LPS relies on specialized macromolecular complexes that span the entire envelope. In this work, we show that Escherichia coli cells are capable of avoiding lysis when the transport of LPS to the OM is compromised, by utilizing LD-transpeptidases (LDTs) to generate 3-3 cross-links in the PG. This PG remodeling program relies mainly on the activities of the stress response LDT, LdtD, together with the major PG synthase PBP1B, its cognate activator LpoB, and the carboxypeptidase PBP6a. Our data support a model according to which these proteins cooperate to strengthen the PG in response to defective OM synthesis. IMPORTANCE In Gram-negative bacteria, the outer membrane protects the cell against many toxic molecules, and the peptidoglycan layer provides protection against osmotic challenges, allowing bacterial cells to survive in changing environments. Maintaining cell envelope integrity is therefore a question of life or death for a bacterial cell. Here we show that Escherichia coli cells activate the LD-transpeptidase LdtD to introduce 3-3 cross-links in the peptidoglycan layer when the integrity of the outer membrane is compromised, and this response is required to avoid cell lysis. This peptidoglycan remodeling program is a strategy to increase the overall robustness of the bacterial cell envelope in response to defects in the outer membrane.

Structural and Functional Variation in Outer Membrane Polysaccharide Export (OPX) Proteins from the Two Major Capsule Assembly Pathways Present in Escherichia coli

“…The CPS export machinery must pass through the PG layer without compromising the essential integrity of the cell wall. The complexity of these systems and the importance of maintaining the PG barrier has recently been highlighted in the context of LPS translocation, where conditional depletion of the translocation machinery immediately activates a PG-editing enzyme complex (37). The nature of the interaction between OPX proteins and PG is unknown, but the noncovalent interaction of the ␤-barrel OM protein OmpA with PG has been studied in detail and residues have been identified that are important for PG association in OmpA and other PG-associated proteins (29,38).…”

pH-dependent activation of cytokinesis modulates Escherichia coli cell size

“…FtsN mutants defective for either domain fail to localize to the cytokinetic ring or to reduce size when overexpressed (Figs 4 and 5). Apart from promoting cytokinesis, FtsN also activates PBP1b [64,74,75], a nonessential cell wall synthesis enzyme speculated to play a role in cell wall repair during normal growth of the E. coli peptidoglycan sacculus [46,[76][77][78]. Midcell enrichment of FtsN in acidic media may also augment PBP1b activity at the septum and thus direct it to the region with the highest rates of peptidoglycan synthesis.…”

Outer membrane lipoprotein NlpI scaffolds peptidoglycan hydrolases within multi‐enzyme complexes in Escherichia coli

“…It has been previously hypothesized that the formation of these complexes enables the cell to coordinate and regulate the activities of various synthetic and hydrolytic PG enzymes in a spatiotemporal manner (Höltje, 1993). Within these complexes, the key bifunctional penicillin-binding protein (PBP) PG synthases are activated by cognate outer membrane (OM)-anchored lipoproteins (Paradis-Bleau et al, 2010;Typas et al, 2010Typas et al, , 2012Dorr et al, 2014;Egan et al, 2014Egan et al, , 2018Greene et al, 2018;Moré et al, 2019) and coordinate their action with another, cell constriction-related protein complex (Gray et al, 2015). However, with the exception of the amidases (Uehara et al, 2010;Yang et al, 2012;Peters et al, 2013;Tsang et al, 2017), it is less clear how Gram-negative bacteria control the activities of their repertoire of hydrolases, i.e.…”