scholarly journals Growth Inhibition by External Potassium of Escherichia coli Lacking PtsN (EIIANtr) Is Caused by Potassium Limitation Mediated by YcgO

2016 ◽  
Vol 198 (13) ◽  
pp. 1868-1882 ◽  
Author(s):  
Ravish Sharma ◽  
Tomohiro Shimada ◽  
Vinod K. Mishra ◽  
Suchitra Upreti ◽  
Abhijit A. Sardesai
Keyword(s):  
Escherichia Coli ◽  
Membrane Protein ◽  
Phosphorylation Site ◽  
Phosphotransferase System ◽  
Inner Membrane ◽  
Content Type ◽  
E Coli ◽  
External Potassium ◽  
Inner Membrane Protein ◽  
External K

ABSTRACTThe absence of PtsN, the terminal phosphoacceptor of the phosphotransferase system comprising PtsP-PtsO-PtsN, inEscherichia coliconfers a potassium-sensitive (Ks) phenotype as the external K+concentration ([K+]e) is increased above 5 mM. A growth-inhibitory increase in intracellular K+content, resulting from hyperactivated Trk-mediated K+uptake, is thought to cause this Ks. We provide evidence that the Ksof the ΔptsNmutant is associated with K+limitation. Accordingly, the moderate Ksdisplayed by the ΔptsNmutant was exacerbated in the absence of the Trk and Kup K+uptake transporters and was associated with reduced cellular K+content. Conversely, overproduction of multiple K+uptake proteins suppressed the Ks. Expression of PtsN variants bearing the H73A, H73D, and H73E substitutions of the phosphorylation site histidine of PtsN complemented the Ks. Absence of the predicted inner membrane protein YcgO (also called CvrA) suppressed the Ks, which was correlated with elevated cellular K+content in the ΔptsNmutant, but the ΔptsNmutation did not alter YcgO levels. Heterologous overexpression ofycgOalso led to Ksthat was associated with reduced cellular K+content, exacerbated by the absence of Trk and Kup and alleviated by overproduction of Kup. Our findings are compatible with a model that postulates that Ksin the ΔptsNmutant occurs due to K+limitation resulting from activation of K+efflux mediated by YcgO, which may be additionally stimulated by [K+]e, implicating a role for PtsN (possibly its dephosphorylated form) as an inhibitor of YcgO activity.IMPORTANCEThis study examines the physiological link between the phosphotransferase system comprising PtsP-PtsO-PtsN and K+ion metabolism inE. coli. Studies on the physiological defect that renders anE. colimutant lacking PtsN to be growth inhibited by external K+indicate that growth impairment results from cellular K+limitation that is mediated by YcgO, a predicted inner membrane protein. Additional observations suggest that dephospho-PtsN may inhibit and external K+may stimulate K+limitation mediated by YcgO. It is speculated that YcgO-mediated K+limitation may be an output of a response to certain stresses, which by modulating the phosphotransfer capacity of the PtsP-PtsO-PtsN phosphorelay leads to growth cessation and stress tolerance.

scholarly journals Tail-Anchored Inner Membrane Protein ElaB Increases Resistance to Stress While Reducing Persistence in Escherichia coli

2017 ◽  
Vol 199 (9) ◽  
Author(s):  
Yunxue Guo ◽  
Xiaoxiao Liu ◽  
Baiyuan Li ◽  
Jianyun Yao ◽  
Thomas K. Wood ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Escherichia Coli ◽  
Membrane Proteins ◽  
Membrane Protein ◽  
Stationary Phase ◽  
Transmembrane Domain ◽  
Inner Membrane ◽  
Content Type ◽  
E Coli ◽  
Inner Membrane Protein

ABSTRACT Host-associated bacteria, such as Escherichia coli, often encounter various host-related stresses, such as nutritional deprivation, oxidative stress, and temperature shifts. There is growing interest in searching for small endogenous proteins that mediate stress responses. Here, we characterized the small C-tail-anchored inner membrane protein ElaB in E. coli. ElaB belongs to a class of tail-anchored inner membrane proteins with a C-terminal transmembrane domain but lacking an N-terminal signal sequence for membrane targeting. Proteins from this family have been shown to play vital roles, such as in membrane trafficking and apoptosis, in eukaryotes; however, their role in prokaryotes is largely unexplored. Here, we found that the transcription of elaB is induced in the stationary phase in E. coli and stationary-phase sigma factor RpoS regulates elaB transcription by binding to the promoter of elaB. Moreover, ElaB protects cells against oxidative stress and heat shock stress. However, unlike membrane peptide toxins TisB and GhoT, ElaB does not lead to cell death, and the deletion of elaB greatly increases persister cell formation. Therefore, we demonstrate that disruption of C-tail-anchored inner membrane proteins can reduce stress resistance; it can also lead to deleterious effects, such as increased persistence, in E. coli. IMPORTANCE Escherichia coli synthesizes dozens of poorly understood small membrane proteins containing a predicted transmembrane domain. In this study, we characterized the function of the C-tail-anchored inner membrane protein ElaB in E. coli. ElaB increases resistance to oxidative stress and heat stress, while inactivation of ElaB leads to high persister cell formation. We also demonstrated that the transcription of elaB is under the direct regulation of stationary-phase sigma factor RpoS. Thus, our study reveals that small inner membrane proteins may have important cellular roles during the stress response.

scholarly journals The Escherichia coli Inner Membrane Protein YhiM is Necessary for Efficient Attachment of Bacteriophage T4

Fine Focus ◽  
2018 ◽  
Vol 4 (1) ◽  
pp. 103-114
Author(s):  
M.A. Evans ◽  
P.T. Spieth ◽  
R.L. Sparks-Thissen
Keyword(s):  
Escherichia Coli ◽  
Membrane Protein ◽  
Bacteriophage T4 ◽  
Inner Membrane ◽  
E Coli ◽  
Obligate Intracellular ◽  
Intracellular Parasites ◽  
Acid Shock ◽  
Inner Membrane Protein

Bacteriophages are obligate intracellular parasites, but many of the cellular proteins involved in replication have not been identified. We have tested the role of the inner membrane protein YhiM in bacteriophage replication. YhiM is a conserved (21) membrane protein in Escherichia coli (E. coli) thought to be localized to the cytoplasmic membrane that is necessary for cell survival under conditions of cell stress, including acid shock, low osmolarity and high temperature. We show here that YhiM is necessary for replication of the bacteriophage T4. It also plays a modest role in the replication of T1, T3, and T5 but it does not play a role in the replication of ΦX174. Our data indicated that no replication of T4 occurs in cells missing YhiM. This block in infection is due to a block in attachment of the virus to the cell surface.

scholarly journals Genetic analysis of the role of the conserved inner membrane protein CvpA in EHEC resistance to deoxycholate

2020 ◽  
Author(s):  
Alyson R. Warr ◽  
Rachel T. Giorgio ◽  
Matthew K. Waldor
Keyword(s):  
Stress Response ◽  
Membrane Protein ◽  
Bile Salt ◽  
Cell Envelope ◽  
Enteric Bacteria ◽  
Enteric Pathogens ◽  
Inner Membrane ◽  
Bacterial Phyla ◽  
E Coli ◽  
Inner Membrane Protein

The function of cvpA, a bacterial gene predicted to encode an inner membrane protein, is largely unknown. Early studies in E. coli linked cvpA to Colicin V secretion and recent work revealed that it is required for robust intestinal colonization by diverse enteric pathogens. In enterohemorrhagic E. coli (EHEC), cvpA is required for resistance to the bile salt deoxycholate (DOC). Here, we carried out genome-scale transposon-insertion mutagenesis and spontaneous suppressor analysis to uncover cvpA’s genetic interactions and identify common pathways that rescue the sensitivity of a ΔcvpA EHEC mutant to DOC. These screens demonstrated that mutations predicted to activate the σE-mediated extracytoplasmic stress response bypass the ΔcvpA mutant’s susceptibility to DOC. Consistent with this idea, we found that deletions in rseA and msbB and direct overexpression of rpoE restored DOC resistance to the ΔcvpA mutant. Analysis of the distribution of CvpA homologs revealed that this inner membrane protein is conserved across diverse bacterial phyla, in both enteric and non-enteric bacteria that are not exposed to bile. Together, our findings suggest that CvpA plays a role in cell envelope homeostasis in response to DOC and similar stress stimuli in diverse bacterial species. IMPORTANCE Several enteric pathogens, including Enterohemorrhagic E. coli (EHEC), require CvpA to robustly colonize the intestine. This inner membrane protein is also important for secretion of a colicin and EHEC resistance to the bile salt deoxycholate (DOC), but its function is unknown. Genetic analyses carried out here showed that activation of the σE-mediated extracytoplasmic stress response restored the resistance of a cvpA mutant to DOC, suggesting that CvpA plays a role in cell envelope homeostasis. The conservation of CvpA across diverse bacterial phyla suggests that this membrane protein facilitates cell envelope homeostasis in response to varied cell envelope perturbations.

scholarly journals YieJ (CbrC) Mediates CreBC-Dependent Colicin E2 Tolerance in Escherichia coli

2010 ◽  
Vol 192 (13) ◽  
pp. 3329-3336 ◽  
Author(s):  
S. James L. Cariss ◽  
Chrystala Constantinidou ◽  
Mala D. Patel ◽  
Yuiko Takebayashi ◽  
Jon L. Hobman ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Membrane Protein ◽  
Microarray Analysis ◽  
Gene Knockout ◽  
Inner Membrane ◽  
Two Component ◽  
Inner Membrane Protein ◽  
Colicin E2 ◽  
K 12

ABSTRACT Colicin E2-tolerant (known as Cet2) Escherichia coli K-12 mutants overproduce an inner membrane protein, CreD, which is believed to cause the Cet2 phenotype. Here, we show that overproduction of CreD in a Cet2 strain results from hyperactivation of the CreBC two-component regulator, but CreD overproduction is not responsible for the Cet2 phenotype. Through microarray analysis and gene knockout and overexpression studies, we show that overexpression of another CreBC-regulated gene, yieJ (also known as cbrC), causes the Cet2 phenotype.

scholarly journals Characterization of YmgF, a 72-Residue Inner Membrane Protein That Associates with the Escherichia coli Cell Division Machinery

2008 ◽  
Vol 191 (1) ◽  
pp. 333-346 ◽  
Author(s):  
Gouzel Karimova ◽  
Carine Robichon ◽  
Daniel Ladant
Keyword(s):  
Escherichia Coli ◽  
Cell Division ◽  
Membrane Protein ◽  
Integral Membrane Protein ◽  
Dependent Manner ◽  
E Coli ◽  
Division Site ◽  
Inner Membrane Protein ◽  
Two Hybrid

ABSTRACT Formation of the Escherichia coli division septum is catalyzed by a number of essential proteins (named Fts) that assemble into a ring-like structure at the future division site. Many of these Fts proteins are intrinsic transmembrane proteins whose functions are largely unknown. In the present study, we attempted to identify a novel putative component(s) of the E. coli cell division machinery by searching for proteins that could interact with known Fts proteins. To do that, we used a bacterial two-hybrid system based on interaction-mediated reconstitution of a cyclic AMP (cAMP) signaling cascade to perform a library screening in order to find putative partners of E. coli cell division protein FtsL. Here we report the characterization of YmgF, a 72-residue integral membrane protein of unknown function that was found to associate with many E. coli cell division proteins and to localize to the E. coli division septum in an FtsZ-, FtsA-, FtsQ-, and FtsN-dependent manner. Although YmgF was previously shown to be not essential for cell viability, we found that when overexpressed, YmgF was able to overcome the thermosensitive phenotype of the ftsQ1(Ts) mutation and restore its viability under low-osmolarity conditions. Our results suggest that YmgF might be a novel component of the E. coli cell division machinery.

scholarly journals Repression of the Inner Membrane Lipoprotein NlpA by Rns in Enterotoxigenic Escherichia coli

2006 ◽  
Vol 189 (5) ◽  
pp. 1627-1632 ◽  
Author(s):  
Maria D. Bodero ◽  
M. Carolina Pilonieta ◽  
George P. Munson
Keyword(s):  
Escherichia Coli ◽  
Outer Membrane ◽  
Membrane Vesicles ◽  
Outer Membrane Vesicles ◽  
Enterotoxigenic Escherichia Coli ◽  
Start Codon ◽  
Inner Membrane ◽  
E Coli ◽  
Inner Membrane Protein

ABSTRACT The expression of the inner membrane protein NlpA is repressed by the enterotoxigenic Escherichia coli (ETEC) virulence regulator Rns, a member of the AraC/XylS family. The Rns homologs CfaD from ETEC and AggR from enteroaggregative E. coli also repress expression of nlpA. In vitro DNase I and potassium permanganate footprinting revealed that Rns binds to a site overlapping the start codon of nlpA, preventing RNA polymerase from forming an open complex at nlpAp. A second Rns binding site between positions −152 and −195 relative to the nlpA transcription start site is not required for repression. NlpA is not essential for growth of E. coli under laboratory conditions, but it does contribute to the biogenesis of outer membrane vesicles. As outer membrane vesicles have been shown to contain ETEC heat-labile toxin, the repression of nlpA may be an indirect mechanism through which the virulence regulators Rns and CfaD limit the release of toxin.

scholarly journals Escherichia coli YqjA, a Member of the Conserved DedA/Tvp38 Membrane Protein Family, Is a Putative Osmosensing Transporter Required for Growth at Alkaline pH

2015 ◽  
Vol 197 (14) ◽  
pp. 2292-2300 ◽  
Author(s):  
Sujeet Kumar ◽  
William T. Doerrler
Keyword(s):  
Escherichia Coli ◽  
Membrane Protein ◽  
Amino Acid Identity ◽  
Protein Family ◽  
Cytoplasmic Ph ◽  
Alkaline Ph ◽  
Ph Homeostasis ◽  
Content Type ◽  
E Coli ◽  
Alkaline Conditions

ABSTRACTThe ability to persist and grow under alkaline conditions is an important characteristic of many bacteria. In order to survive at alkaline pH,Escherichia colimust maintain a stable cytoplasmic pH of about 7.6. Membrane cation/proton antiporters play a major role in alkaline pH homeostasis by catalyzing active inward proton transport. The DedA/Tvp38 family is a highly conserved membrane protein family of unknown function present in most sequenced genomes. YqjA and YghB are members of theE. coliDedA family with 62% amino acid identity and partially redundant functions. We have shown thatE. coliwith ΔyqjAand ΔyghBmutations cannot properly maintain the proton motive force (PMF) and is compromised in PMF-dependent drug efflux and other PMF-dependent functions. Furthermore, the functions of YqjA and YghB are dependent upon membrane-embedded acidic amino acids, a hallmark of several families of proton-dependent transporters. Here, we show that the ΔyqjAmutant (but not ΔyghB) cannot grow under alkaline conditions (ranging from pH 8.5 to 9.5), unlike the parentE. coli. Overexpression ofyqjArestores growth at alkaline pH, but only when more than ∼100 mM sodium or potassium is present in the growth medium. Increasing the osmotic pressure by the addition of sucrose enhances the ability of YqjA to support growth under alkaline conditions in the presence of low salt concentrations, consistent with YqjA functioning as an osmosensor. We suggest that YqjA possesses proton-dependent transport activity that is stimulated by osmolarity and that it plays a significant role in the survival ofE. coliat alkaline pH.IMPORTANCEThe ability to survive under alkaline conditions is important for many species of bacteria.Escherichia colican grow at pH 5.5 to 9.5 while maintaining a constant cytoplasmic pH of about 7.6. Under alkaline conditions, bacteria rely upon proton-dependent transporters to maintain a constant cytoplasmic pH. The DedA/Tvp38 protein family is a highly conserved but poorly characterized family of membrane proteins. Here, we show that the DedA/Tvp38 protein YqjA is critical forE. colito survive at pH 8.5 to 9.5. YqjA requires sodium and potassium for this function. At low cation concentrations, osmolytes, including sucrose, can facilitate rescue ofE. coligrowth by YqjA at high pH. These data are consistent with YqjA functioning as an osmosensing cation-dependent proton transporter.

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