Abstract
ABSTRACTThe ability of Gram-negative bacteria to carefully modulate outer membrane (OM) composition is essential to their survival. However, the asymmetric and heterogeneous structure of the Gram-negative OM poses unique challenges to the cell’s successful adaption to rapid environmental transitions. Although mechanisms to recycle and degrade OM phospholipid material exist, there is no known mechanism by which to remove unfavorable lipopolysaccharide (LPS) glycoforms, except slow dilution through cell growth. As all Gram-negative bacteria constitutively shed OM vesicles (OMVs), we propose that cells may utilize OMV formation as a way to selectively remove environmentally disadvantageous LPS species. We examined the native kinetics of OM composition during physiologically relevant environmental changes in Salmonella enterica, a well-characterized model system for activation of PhoP/Q and PmrA/B two-component systems (TCSs). In response to acidic pH, toxic metals, antimicrobial peptides, and lack of divalent cations, these TCSs modify the LPS lipid A and core, lengthen the O antigen, and upregulate specific OM proteins. An environmental change to PhoP/Q- and PmrA/B-activating conditions simultaneously induced the addition of modified species of LPS to the OM, downregulation of previously dominant species of LPS, greater OMV production, and increased OMV diameter. Comparison of the relative abundance of lipid A species present in the OM and the newly budded OMVs following two sets of rapid environmental shifts revealed the retention of lipid A species with modified phosphate moieties in the OM concomitant with the selective loss of palmitoylated species via vesiculation following exposure to moderately acidic environmental conditions.
Highlights
The ability of Gram-negative bacteria to carefully modulate outer membrane (OM) composition is essential to their survival
In order to investigate the role of OM vesicles (OMVs) in the mediation and maintenance of OM remodeling, it was necessary to develop a protocol that would allow us to examine wild-type cells that are induced to accomplish a significant, measurable OM change in response to a physiologically relevant environmental shift but have a minimal opportunity to remodel their membrane via dilution upon cell growth and division
We further decided to focus on LPS since there are few known OM-localized enzymes that are known to act on this component after it has reached the OM
Summary
The ability of Gram-negative bacteria to carefully modulate outer membrane (OM) composition is essential to their survival. In response to a mildly acidic pH, a high ferric iron level, and toxic aluminum concentrations, the PmrA/B TCS activates genes responsible for modulation of O-antigen length (wzzfepE, wzzST), LPS core decorations (pmrG, cptA), and lipid A modifications (pmrC, pmrR, pbgP) [5]. These modifications are known to be critical for Salmo-. Once generated, sufficient levels of modified LPS species must be retained in order for the cell to derive maximal benefit from the energy-consuming biosynthetic process
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