LapB (YciM) orchestrates protein–protein interactions at the interface of lipopolysaccharide and phospholipid biosynthesis

Möller, Anna-Maria; Brückner, Simon; Tilg, Lea‐Janina; Kutscher, Blanka; Nowaczyk, Marc M.; Narberhaus, Franz

doi:10.1111/mmi.15005

Cited by 11 publications

(12 citation statements)

References 68 publications

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“…E. coli LapB interacts with other critical enzymes for LPS and GPL biosynthesis ( 25 , 28 ). However, the LapB regions and residues that mediate these interactions are not known.…”

Section: Resultsmentioning

confidence: 99%

“…1B ). In the “dissociation model,” PbgA binds these LPS molecules causing LapB to dissociate from PbgA, which frees LapB to bind LpxC and FtsH and enhance LpxC proteolysis ( 26 – 28 , 34 , 36 ). By contrast, the “constitutive-complex model” predicts that the PbgA-LapB complex does not dissociate when nutrients become limiting.…”

Section: Introductionmentioning

confidence: 99%

“…Likewise, as shown recently for E. coli , PbgA and LpxC interact in S . Typhimurium ( 28 ). In summary, PbgA-LapB is a keystone regulator of enterobacterial cell envelope biogenesis that impinges on LpxC in a manner that could inform new and existing antimicrobials that target LPS biosynthesis enzymes.…”

Section: Introductionmentioning

confidence: 99%

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Signaling through the Salmonella PbgA-LapB regulatory complex activates LpxC proteolysis and limits lipopolysaccharide biogenesis during stationary-phase growth

Mettlach,

Cian,

Chakraborty

et al. 2024

J Bacteriol

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Salmonella enterica serovar Typhimurium ( S . Typhimurium) controls lipopolysaccharide (LPS) biosynthesis by regulating proteolysis of LpxC, the rate-limiting enzyme and target of preclinical antibiotics. PbgA/YejM/LapC regulates LpxC levels and controls outer membrane (OM) LPS composition at the log-to-stationary phase transition. Suppressor substitutions in L PS a ssembly p rotein B (LapB/YciM) rescue the LPS and OM integrity defects of pbgA -mutant S . Typhimurium. We hypothesized that PbgA regulates LpxC proteolysis by controlling LapB’s ability to bind LpxC as a function of the growth phase. According to existing models, when nutrients are abundant, PbgA binds and restricts LapB from interacting with LpxC and FtsH, which limits LpxC proteolysis. However, when nutrients are limited, there is debate whether LapB dissociates from PbgA to bind LpxC and FtsH to enhance degradation. We sought to examine these models and investigate how the structure of LapB enables salmonellae to control LpxC proteolysis and LPS biosynthesis. Salmonellae increase LapB levels during the stationary phase to promote LpxC degradation, which limits lipid A-core production and increases their survival. The deletion of lapB , resulting in unregulated lipid A-core production and LpxC overabundance, leads to bacterial growth retardation. Tetratricopeptide repeats near the cytosol-inner membrane interface are sufficient for LapB to bind LpxC, and remarkably, LapB and PbgA interact in both growth phases, yet LpxC only associates with LapB in the stationary phase. Our findings support that PbgA-LapB exists as a constitutive complex in S . Typhimurium, which differentially binds LpxC to control LpxC proteolysis and limit lipid A-core biosynthesis in response to changes in the environment. IMPORTANCE Antimicrobial resistance has been a costly setback for human health and agriculture. Continued pursuit of new antibiotics and targets is imperative, and an improved understanding of existing ones is necessary. LpxC is an essential target of preclinical trial antibiotics that can eliminate multidrug-resistant Gram-negative bacterial infections. LapB is a natural LpxC inhibitor that targets LpxC for degradation and limits lipopolysaccharide production in Enterobacteriaceae. Contrary to some studies, findings herein support that LapB remains in complex instead of dissociating from its presumed negative regulator, PbgA/YejM/LapC, under conditions where LpxC proteolysis is enhanced. Advanced comprehension of this critical protein-lipid signaling network will lead to future development and refinement of small molecules that can specifically interfere.

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“…E. coli LapB interacts with other critical enzymes for LPS and GPL biosynthesis ( 25 , 28 ). However, the LapB regions and residues that mediate these interactions are not known.…”

Section: Resultsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 1 more Smart Citation

Signaling through the Salmonella PbgA-LapB regulatory complex activates LpxC proteolysis and limits lipopolysaccharide biogenesis during stationary-phase growth

Mettlach,

Cian,

Chakraborty

et al. 2024

J Bacteriol

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“…Residual cell debris were removed by filtration of the supernatant (0,2 μm filter). The supernatant was finally loaded on a Ni‐NTA column and purified as previously described (Möller et al., 2023).…”

Section: Methodsmentioning

confidence: 99%

The LysR‐type transcription factor LsrB regulates beta‐lactam resistance in Agrobacterium tumefaciens

Schmidt,

Remme,

Eisfeld

et al. 2023

Molecular Microbiology

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Agrobacterium tumefaciens is a plant pathogen, broadly known as the causal agent of the crown gall disease. The soil bacterium is naturally resistant to beta‐lactam antibiotics by utilizing the inducible beta‐lactamase AmpC. Our picture on the condition‐dependent regulation of ampC expression is incomplete. A known regulator is AmpR controlling the transcription of ampC in response to unrecycled muropeptides as a signal for cell wall stress. In our study, we uncovered the global transcriptional regulator LsrB as a critical player acting upstream of AmpR. Deletion of lsrB led to severe ampicillin and penicillin sensitivity, which could be restored to wild‐type levels by lsrB complementation. By transcriptome profiling via RNA‐Seq and qRT‐PCR and by electrophoretic mobility shift assays, we show that ampD coding for an anhydroamidase involved in peptidoglycan recycling is under direct negative control by LsrB. Controlling AmpD levels by the LysR‐type regulator in turn impacts the cytoplasmic concentration of cell wall degradation products and thereby the AmpR‐mediated regulation of ampC. Our results substantially expand the existing model of inducible beta‐lactam resistance in A. tumefaciens by establishing LsrB as higher‐level transcriptional regulator.

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“…LpxC levels went up and stayed stable upon protein synthesis inhibition indicating that inhibitor-bound enzyme is (i) inactive because LPS levels remained low and (ii) not degraded by the FtsH protease. The latter can probably be explained by the inaccessibility of recognition motifs in LpxC (14,47), the inability to interact with the machinery that delivers LpxC to FtsH (48)(49)(50), or changes in metabolites, e.g. lipid A disaccharide, acyl-CoA or periplasmic LPS intermediates, affecting LpxC stability (19,51,52).…”

Section: Similar Yet Different Responses Of E Coli To Lpxc Inhibitionmentioning

confidence: 99%

Same same but different – The global response ofEscherichia colito five different LpxC inhibitors

Möller,

Vázquez-Hernández,

Kutscher

et al. 2023

Preprint

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A promising but yet clinically unexploited antibiotic target in difficult-to-treat Gram-negative bacteria is LpxC, the key enzyme in the biosynthesis of lipopolysaccharides (LPS), which are the major constituents of the outer membrane. To gain insights into the mode of action of five different LpxC inhibitors, we conducted a comparative phenotypic and proteomic analysis. All five compounds bound to purified LpxC from Escherichia coli. Treatment of E. coli with these compounds changed the cell shape and stabilized LpxC suggesting that the FtsH-mediated turnover is impaired. LpxC inhibition sensitized E. coli to the cell wall antibiotic vancomycin, which typically does not cross the outer membrane. Four of the five compounds led to an accumulation of lyso-PE, a cleavage product of phosphatidylethanolamine (PE), generated by the phospholipase PldA. The combined results suggested an imbalance in phospholipid (PL) and LPS biosynthesis, which was corroborated by the global proteome response to treatment with the LpxC inhibitors. Apart from LpxC itself, FabA and FabB responsible for the biosynthesis of unsaturated fatty acids, were consistently upregulated. Our work also shows that antibiotics targeting the same enzyme do not necessarily elicit identical cellular responses. Compound-specific marker proteins belonged to different functional categories, like stress responses, nucleotide or amino acid metabolism and quorum sensing. These findings provide new insights into common and distinct cellular defense mechanisms against LpxC inhibition. Moreover, they support a delicate balance between LPS and PL biosynthesis with great potential as point of attack for antimicrobial intervention.

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LapB (YciM) orchestrates protein–protein interactions at the interface of lipopolysaccharide and phospholipid biosynthesis

Cited by 11 publications

References 68 publications

Signaling through the Salmonella PbgA-LapB regulatory complex activates LpxC proteolysis and limits lipopolysaccharide biogenesis during stationary-phase growth

Signaling through the Salmonella PbgA-LapB regulatory complex activates LpxC proteolysis and limits lipopolysaccharide biogenesis during stationary-phase growth

The LysR‐type transcription factor LsrB regulates beta‐lactam resistance in Agrobacterium tumefaciens

Same same but different – The global response ofEscherichia colito five different LpxC inhibitors

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