Dataset Information

Swarmer Cell Development of the Bacterium Proteus mirabilis Requires the Conserved Enterobacterial Common Antigen Biosynthesis Gene rffG.

ABSTRACT: Individual cells of the bacterium Proteus mirabilis can elongate up to 40-fold on surfaces before engaging in a cooperative surface-based motility termed swarming. How cells regulate this dramatic morphological remodeling remains an open question. In this paper, we move forward the understanding of this regulation by demonstrating that P. mirabilis requires the gene rffG for swarmer cell elongation and subsequent swarm motility. The rffG gene encodes a protein homologous to the dTDP-glucose 4,6-dehydratase protein of Escherichia coli, which contributes to enterobacterial common antigen biosynthesis. Here, we characterize the rffG gene in P. mirabilis, demonstrating that it is required for the production of large lipopolysaccharide-linked moieties necessary for wild-type cell envelope integrity. We show that the absence of the rffG gene induces several stress response pathways, including those controlled by the transcriptional regulators RpoS, CaiF, and RcsB. We further show that in rffG-deficient cells, the suppression of the Rcs phosphorelay, via loss of RcsB, is sufficient to induce cell elongation and swarm motility. However, the loss of RcsB does not rescue cell envelope integrity defects and instead results in abnormally shaped cells, including cells producing more than two poles. We conclude that an RcsB-mediated response acts to suppress the emergence of shape defects in cell envelope-compromised cells, suggesting an additional role for RcsB in maintaining cell morphology under stress conditions. We further propose that the composition of the cell envelope acts as a checkpoint before cells initiate swarmer cell elongation and motility.IMPORTANCEProteus mirabilis swarm motility has been implicated in pathogenesis. We have found that cells deploy multiple uncharacterized strategies to handle cell envelope stress beyond the Rcs phosphorelay when attempting to engage in swarm motility. While RcsB is known to directly inhibit the master transcriptional regulator for swarming, we have shown an additional role for RcsB in protecting cell morphology. These data support a growing appreciation that the Rcs phosphorelay is a multifunctional regulator of cell morphology in addition to its role in microbial stress responses. These data also strengthen the paradigm that outer membrane composition is a crucial checkpoint for modulating entry into swarm motility. Furthermore, the rffG-dependent moieties provide a novel attractive target for potential antimicrobials.

SUBMITTER: Little K

PROVIDER: S-EPMC6112005 | biostudies-literature | 2018 Sep

REPOSITORIES: biostudies-literature

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Swarmer Cell Development of the Bacterium Proteus mirabilis Requires the Conserved Enterobacterial Common Antigen Biosynthesis Gene rffG.

Little Kristin K Tipping Murray J MJ Gibbs Karine A KA

Journal of bacteriology 20180824 18

Individual cells of the bacterium Proteus mirabilis can elongate up to 40-fold on surfaces before engaging in a cooperative surface-based motility termed swarming. How cells regulate this dramatic morphological remodeling remains an open question. In this paper, we move forward the understanding of this regulation by demonstrating that P. mirabilis requires the gene rffG for swarmer cell elongation and subsequent swarm motility. The rffG gene encodes a protein homolog ...[more]

PMID: 29967121

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Project description:The Gram-negative cell envelope is a complex structure delineating the cell from its environment. Recently, we found that enterobacterial common antigen (ECA) plays a role maintaining the outer membrane (OM) permeability barrier, which excludes toxic molecules including many antibiotics. ECA is a conserved carbohydrate found throughout Enterobacterales (e.g., Salmonella, Klebsiella, and Yersinia). There are two OM forms of ECA (phosphoglyceride-linked ECAPG and lipopolysaccharide-linked ECALPS) and one periplasmic form of ECA (cyclic ECACYC). ECAPG, found in the outer leaflet of the OM, consists of a linear ECA oligomer attached to phosphoglyceride through a phosphodiester linkage. The process through which ECAPG is produced from polymerized ECA is unknown. Therefore, we set out to identify genes interacting genetically with ECAPG biosynthesis in Escherichia coli K-12 using the competition between ECA and peptidoglycan biosynthesis. Through transposon-directed insertion sequencing, we identified an interaction between elyC and ECAPG biosynthesis. ElyC is an inner membrane protein previously shown to alter peptidoglycan biosynthesis rates. We found ΔelyC was lethal specifically in strains producing ECAPG without other ECA forms, suggesting ECAPG biosynthesis impairment or dysregulation. Further characterization suggested ElyC inhibits ECAPG synthesis in a posttranscriptional manner. Moreover, the full impact of ElyC on ECA levels requires the presence of ECACYC. Our data demonstrate ECACYC can regulate ECAPG synthesis in strains wild type for elyC. Overall, our data demonstrate ElyC and ECACYC act in a novel pathway that regulates the production of ECAPG, supporting a model in which ElyC provides feedback regulation of ECAPG production based on the periplasmic levels of ECACYC. IMPORTANCE Enterobacterial common antigen (ECA) is a conserved polysaccharide present on the surface of the outer membrane (OM) and in the periplasm of the many pathogenic bacteria belonging to Enterobacterales, including Klebsiella pneumoniae, Salmonella enterica, and Yersinia pestis. As the OM is a permeability barrier that excludes many antibiotics, synthesis pathways for OM molecules are promising targets for antimicrobial discovery. Here, we elucidated, in E. coli K-12, a new pathway for the regulation of biosynthesis of one cell surface form of ECA, ECAPG. In this pathway, an inner membrane protein, ElyC, and the periplasmic form of ECA, ECACYC, genetically interact to inhibit the synthesis of ECAPG, potentially through feedback regulation based on ECACYC levels. This is the first insight into the pathway responsible for synthesis of ECAPG and represents a potential target for weakening the OM permeability barrier. Furthermore, this pathway provides a tool for experimental manipulation of ECAPG levels.

Dataset Information

Swarmer Cell Development of the Bacterium Proteus mirabilis Requires the Conserved Enterobacterial Common Antigen Biosynthesis Gene rffG.

Publications

Swarmer Cell Development of the Bacterium Proteus mirabilis Requires the Conserved Enterobacterial Common Antigen Biosynthesis Gene <i>rffG</i>.

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