Assembly of Lipopolysaccharide in Escherichia coli Requires the Essential LapB Heat Shock Protein

Klein, Gracjana; Kobylak, Natalia; Lindner, Buko; Stupak, Anna; Raina, Satish

doi:10.1074/jbc.m113.539494

Cited by 94 publications

(284 citation statements)

References 67 publications

(110 reference statements)

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“…Previously, we showed that the balanced biosynthesis of LPS requires the essential LapB protein and in its absence transcription of the rpoE gene is significantly induced (9). ⌬lapB mutants synthesize excess LPS with a significant proportion of LPS comprised of precursor forms and also have a dysfunctional Lpt translocation system (9).…”

Section: Defects In the Lps Assembly Also Induce The Rpoep3 Promotermentioning

confidence: 99%

“…Thus, one might miss genes whose products require the expression of whole operons or regulatory sRNAs. To overcome such limitations, a previously described complete genomic library in a mediumcopy p15A vector (9,35) was introduced to identify clones that exhibit a Lac up or down phenotype. These multicopy approaches identified qseG and several genes encoding lipoproteins (csgG, pgaB, spr, yhdV, yceB, yddW, and yghB), which, when overexpressed, increase the rpoEP-lacZ activity (Tables 1 and 2).…”

Section: Tablementioning

confidence: 99%

“…Subsequently, we showed that strains synthesizing the minimal LPS structure composed of either Kdo 2 -lipid IV A or only free lipid IV A exhibited hyperelevated levels of the RpoE activity (6). The activity of RpoE is highly induced when the LPS assembly is severely compromised or when there is an imbalance between LPS and phospholipids (9). Lack of many LPS core biosynthetic genes also leads to defects in survival at high temperatures (5,6,10).…”

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confidence: 99%

“…The synthesis of such LPS glycoforms is controlled by the RpoEregulated RybB and EptB (19). RpoE also positively regulates transcription of micA and slrA (micL) sRNAs (9,20,21). MicA negatively regulates the PhoP expression by direct translational inhibition of the phoP mRNA (22), thus further linking the PhoP/Q two-component system, known to regulate LPS nonstoichiometric alterations, with the RpoE control.…”

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confidence: 99%

“…Bacteriophage P1 or T4 lysates were prepared on individual Lac up candidates and transduced back into SR4245 or SR7197. The position of Tn10 was determined by the inverse PCR with nested primers and sequenced using the Tn10 primer as described previously (9).…”

mentioning

confidence: 99%

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Multiple Transcriptional Factors Regulate Transcription of the rpoE Gene in Escherichia coli under Different Growth Conditions and When the Lipopolysaccharide Biosynthesis Is Defective

Klein

Stupak

Biernacka

et al. 2016

Journal of Biological Chemistry

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107

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The RpoE factor is essential for the viability of Escherichia coli. RpoE regulates extracytoplasmic functions including lipopolysaccharide (LPS) translocation and some of its non-stoichiometric modifications. Transcription of the rpoE gene is positively autoregulated by E E and by unknown mechanisms that control the expression of its distally located promoter(s). Mapping of 5 ends of rpoE mRNA identified five new transcriptional initiation sites (P1 to P5) located distal to E E -regulated promoter. These promoters are activated in response to unique signals. Of these P2, P3, and P4 defined major promoters, recognized by RpoN, RpoD, and RpoS factors, respectively. Isolation of trans-acting factors, in vitro transcriptional and gel retardation assays revealed that the RpoN-recognized P2 promoter is positively regulated by a QseE/F two-component system and NtrC activator, whereas the RpoD-regulated P3 promoter is positively regulated by a Rcs system in response to defects in LPS core biosynthesis, overproduction of certain lipoproteins, and the global regulator CRP. Strains synthesizing Kdo 2 -LA LPS caused up to 7-fold increase in the rpoEP3 activity, which was abrogated in ⌬(waaC rcsB). Overexpression of a novel 73-nucleotide sRNA rirA (RfaH interacting RNA) generated by the processing of 5 UTR of the waaQ mRNA induces the rpoEP3 promoter activity concomitant with a decrease in LPS content and defects in the O-antigen incorporation. In the presence of RNA polymerase, RirA binds LPS regulator RfaH known to prevent premature transcriptional termination of waaQ and rfb operons. RirA in excess could titrate out RfaH causing LPS defects and the activation of rpoE transcription.The cell envelope of Gram-negative bacteria, including Escherichia coli, contains two distinct membranes, an inner (IM) 3 and an outer (OM) membrane separated by the periplasm, a hydrophilic compartment that includes a layer of peptidoglycan. The OM is an asymmetric lipid bilayer with phospholipids forming the inner leaflet and LPS forming the outer leaflet. LPSs are highly heterogeneous in composition. However, they share a common architecture composed of a membrane-anchored phosphorylated and acylated ␤(136)-linked GlcN disaccharide, termed lipid A, to which a carbohydrate moiety of varying size is attached (1, 2).During the analyses of signals that induce the RpoE-dependent extracytoplasmic stress response, we showed that strains synthesizing heptoseless LPS due to the absence of either GmhD (RfaD/HtrM) or WaaC exhibited a constitutive induction of RpoE (3-6). Such strains also display constitutive synthesis of exopolysaccharide that is regulated by the Rcs twocomponent system (5,7,8). Subsequently, we showed that strains synthesizing the minimal LPS structure composed of either Kdo 2 -lipid IV A or only free lipid IV A exhibited hyperelevated levels of the RpoE activity (6). The activity of RpoE is highly induced when the LPS assembly is severely compromised or when there is an imbalance between LPS and phospholipids (9). Lack of many LPS co...

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Section: Defects In the Lps Assembly Also Induce The Rpoep3 Promotermentioning

confidence: 99%

Section: Tablementioning

confidence: 99%

mentioning

confidence: 99%

mentioning

confidence: 99%

mentioning

confidence: 99%

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Multiple Transcriptional Factors Regulate Transcription of the rpoE Gene in Escherichia coli under Different Growth Conditions and When the Lipopolysaccharide Biosynthesis Is Defective

Klein

Stupak

Biernacka

et al. 2016

Journal of Biological Chemistry

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107

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Fat Matters for Bugs: How Lipids and Lipid Modifications Make the Difference in Bacterial Life

Sperandeo

Polissi

Fabiani

2019

Euro J Lipid Sci & Tech

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Lipids are building blocks of biological membranes in the three domains of life and are endowed with several important biological functions. Lipopolysaccharide (LPS) is a peculiar glycolipid that represents the hallmark of the cell envelope of Gram‐negative bacteria, a group comprising several important human pathogens. The presence of LPS in the outer membrane (OM) of Gram‐negative bacteria correlates not only with the intrinsic resistance of this group of bacteria toward several antibiotics currently in use, but also with the worrisome increase in antibiotic resistance that is depleting the arsenal of efficacious molecules to cure bacterial infections. LPS consists of a conserved internal moiety decorated by a more variable distal unit. Several modifications occur at the canonical LPS structure during bacterial infection which are exploited by the microorganism to co‐evolve with the host thus evading its immune system. This viewpoint article discusses the current knowledge on LPS biogenesis and modification systems and their consequences on recognition by immune cells and antimicrobial resistance. It also discusses how knowledge on LPS biogenesis can be exploited to develop molecules able to dismantle the Gram‐negative protective barrier and to interfere with the interaction with the mammalian host. Practical Applications: Antibiotic resistance is a major threat for human health leading to inefficacy of many antibiotics to treat infectious diseases. Resistance to antibiotics causes around 25 000 deaths per year in the European Union alone and 700 000 deaths per year globally. This results in an increase of 1.5 billion euros per year in healthcare costs and productivity losses for illness. Gram‐negative bacteria are intrinsically resistant to many antibiotics, due to their LPS‐coated cell surface that protects them from the environment. LPS is sensed by the host immune system as a signal of bacterial infection, moreover bacteria exploit the complexity of LPS modifications to modulate the host immune response and eventually escape it. A deep understanding of the molecular mechanisms underlying LPS biogenesis, including the physico‐chemical and biological consequences of its lipid modifications, will be instrumental to develop novel antibiotic treatments aimed at dismantling the Gram‐negative bacteria barrier and restoring drug sensitivity. The hallmark of Gram‐negative bacteria is the presence of an asymmetric outer membrane (OM) surrounding the cytoplasmic membrane (inner membrane, IM), whose outer leaflet is made by lipopolysaccharide (LPS). A layer of peptidoglycan (PG) is sandwiched in between. LPS endows the bacteria with increased resistance to many antibiotics and is the first line of interaction with the host cell immune system. Chemical modifications of the canonical LPS structure determine different stimulatory effects of the immune response and allow the bacteria to escape from the killing action of effector molecules such as antimicrobial peptides.

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Lipopolysaccharide Biosynthesis and Transport to the Outer Membrane of Gram-Negative Bacteria

Sperandeo

Martorana

Polissi

2019

Subcellular Biochemistry

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Assembly of Lipopolysaccharide in Escherichia coli Requires the Essential LapB Heat Shock Protein

Cited by 94 publications

References 67 publications

Multiple Transcriptional Factors Regulate Transcription of the rpoE Gene in Escherichia coli under Different Growth Conditions and When the Lipopolysaccharide Biosynthesis Is Defective

Multiple Transcriptional Factors Regulate Transcription of the rpoE Gene in Escherichia coli under Different Growth Conditions and When the Lipopolysaccharide Biosynthesis Is Defective

Fat Matters for Bugs: How Lipids and Lipid Modifications Make the Difference in Bacterial Life

Lipopolysaccharide Biosynthesis and Transport to the Outer Membrane of Gram-Negative Bacteria

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