Inner Membrane Protein YhcB Interacts with RodZ Involved in Cell Shape Maintenance in <i>Escherichia coli</i>

Li, Gaochi; Hamamoto, Kentaro; Kitakawa, Madoka

doi:10.5402/2012/304021

Cited by 15 publications

(26 citation statements)

References 34 publications

(41 reference statements)

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“…The encoded YhcB was initially proposed to be a putative third subunit of cytochrome bd-type ubiquinol oxidase (CydAB) [42,67], but a subsequent study showed that it is dispensable for the assembly and function of CydAB [41]. A recent study showed that the yhcB and rodZ double-deletion was lethal while strains with either single deletion were viable, and the authors suggested that YhcB helps maintain the cell shape by joining RodZ in contributing to cell envelope biogenesis [40]. Here, we report that yhcB deletion cells show severe filamentous phenotypes consistent with the yhcB knockdown phenotype.…”

Section: Viabiltiy (%) Lysis Ratiomentioning

confidence: 99%

“…We used RNA-seq data to predict target mRNAs and applied bioinformatics to the minimal region of SdsR. This combined strategy enabled us to identify the major SdsR mRNA target whose repression led to cell death: yhcB, which encodes an inner membrane protein involved in biogenesis of the cell envelope and the maintenance of cell shape [40][41][42].…”

Section: Introductionmentioning

confidence: 99%

“…SdsR overexpression: vulnerability to lysis, cell death and cell filamentation. The yhcB gene is a thermo-tolerant gene that encodes an inner membrane protein involved in biogenesis of the cell envelope and the maintenance of cell shape[40][41][42]44]. While the nt 34-41 sequence of SdsR can base pair with the yhcB coding sequence at nt + 15 to + 22 relative to the start codon, ARyhcB can make base-pair interactions with the yhcB mRNA sequence at nt −19 to + 1(Figure 8(c)).…”

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confidence: 99%

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The small RNA, SdsR, acts as a novel type of toxin inEscherichia coli

et al. 2018

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Most small noncoding RNAs (sRNAs) are known to base pair with target mRNAs and regulate mRNA stability or translation to trigger various changes in the cell metabolism of Escherichia coli. The SdsR sRNA is expressed specifically during the stationary phase and represses tolC and mutS expression. However, it was not previously known whether the growth-phase-dependent regulation of SdsR is important for cell growth. Here, we ectopically expressed SdsR during the exponential phase and examined cell growth and survival. We found that ectopic expression of SdsR led to a significant and Hfq-dependent cell death with accompanying cell filamentation. This SdsR-driven cell death was alleviated by overexpression of RyeA, an sRNA transcribed on the opposite DNA strand, suggesting that SdsR/RyeA is a novel type of toxin-antitoxin (T/A) system in which both the toxin and the antitoxin are sRNAs. We defined the minimal region required for the SdsR-driven cell death. We also performed RNA-seq analysis and identified 209 genes whose expression levels were altered by more than twofold following pulse expression of ectopic SdsR at exponential phase. Finally, we found that that the observed SdsR-driven cell death was mainly caused by the SdsR-mediated repression of yhcB, which encodes an inner membrane protein.

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Section: Viabiltiy (%) Lysis Ratiomentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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confidence: 99%

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The small RNA, SdsR, acts as a novel type of toxin inEscherichia coli

et al. 2018

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“…Recently, our group showed that SdsR and RyeA act as toxin and antitoxin, respectively 18 . The toxin function of SdsR is mediated by repression of yhcB encoding an inner membrane protein, which is involved in cell envelope biogenesis and cell shape maintenance 25–27 . Since both the toxin and antitoxin are sRNAs, the SdsR/RyeA pair represents a novel type of toxin-antitoxin system.…”

Section: Introductionmentioning

confidence: 99%

Coordinate regulation of the expression of SdsR toxin and its downstream pphA gene by RyeA antitoxin in Escherichia coli

Choi

Park

Kim

et al. 2019

Sci Rep

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In Escherichia coli , SdsR and RyeA, a unique pair of mutually cis -encoded small RNAs (sRNAs), act as toxin and antitoxin, respectively. SdsR and RyeA expression are reciprocally regulated; however, how each regulates the synthesis of the other remains unclear. Here, we characterized the biosynthesis of the two sRNAs during growth and investigated their coordinate regulation using sdsR and ryeA promoter mutant strains. We found that RyeA transcription occurred even upon entry of cells into the stationary phase, but its apparent expression was restricted to exponentially growing cells because of its degradation by SdsR. Likewise, the appearance of SdsR was delayed owing to its RyeA-mediated degradation. We also found that the sdsR promoter was primarily responsible for transcription of the downstream pphA gene encoding a phosphatase and that pphA mRNA was synthesized by transcriptional read-through over the sdsR terminator. Transcription from the σ 70 -dependent ryeA promoter inhibited transcription from the σ S -dependent sdsR promoter through transcriptional interference. This transcriptional inhibition also downregulated pphA expression, but RyeA itself did not downregulate pphA expression.

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“…An appropriate amount of cells from each transformant was suspended in LB medium at an OD 600 of about 0.5 and the β‐galactosidase activities measured to quantify T25‐T18 complementation by the interaction between the proteins encoded. The assay of β‐galactosidase activity was performed according to Miller with a minor modification . Average and SD were calculated from three independent measurements.…”

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confidence: 99%

Characterization of inner membrane protein YciB in Escherichia coli: YciB interacts with cell elongation and division proteins

Badaluddin

Kitakawa

2015

Microbiology and Immunology

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The function of inner membrane protein YciB in Escherichia coli has not been identified. In this study, the membrane topology of the protein that contains five transmembrane domains was clarified. YciB was found to interact with various proteins involved in cell elongation and cell division using a bacterial two-hybrid system. It was also found that the deletion mutant of yciB is susceptible to the low osmolarity. These observations together with previous reports indicate that YciB is involved in synthesis of the cell envelope by interacting with cell elongation and cell division complexes.Key words Escherichia coli, membrane topology, two-hybrid analysis, YciB.Bacterial cell envelopes that consist of an IM and the cell wall are the principal stress-bearing and shape-maintaining elements of cells; their integrity is of critical importance to cell viability. The E. coli genome contains over 800 genes (1) that encode IM proteins; however, many of them have not yet been well characterized and their functions remain unknown. We recently identified that one such gene, yciB, is a gene that is required for normal biofilm formation (2) and interacts genetically with rodZ, a gene that is important for maintaining rodtype morphology (3-5). These findings suggest that YciB has a role in cell envelope synthesis.YciB is predicted to be a multi-pass IM protein with five transmembrane domains (Fig. 1a); however, this prediction has not experimentally been examined. Therefore, we first investigated the membrane topology of YciB using a dual pho-lac reporter system (6). In this system, the color on the indicator plate of E. coli strain DH5a (phoA -lacZ -DM15) carrying the pKTop plasmid shows whether the C-terminus of the protein expressed from the cloned gene resides in the cytoplasm or in the periplasm. According to the predicted structure of YciB, we designed serial mutants with deletions from the Cterminus, as shown in Figure 1a, and cloned each fragment into pKTop. The resultant pKTop clones (Table S1) were introduced into DH5a and the topology of the fusion proteins analyzed. Transformants expressing YciB 147 and YciB 75 fusion proteins exhibited blue color, indicating strong phosphatase but low b-galactosidase activity, which indicates that the C-termini of these positions are localized on the periplasmic side of the inner membrane. In contrast, full length YciB, YciB 118 , YciB 48 fusion proteins showed high b-galactosidase activity but low phosphatase activity (red color), showing that the C-termini of these proteins are located in the cytoplasm (Fig. 1b). We also constructed a pKTop recombinant expressing YciB 42-179 fusion. This fusion protein lacks the N-terminal region that is predicted to be periplasmic and the first transmembrane domain. We found that the C-terminus of this mutant is cytoplasmic, as is the full length YciB. Similarly, YciB 42-147 fusion that we deleted at both the N-and C-termini gave the same topological result (blue color) as YciB 147 . These results suggest that neither the N-terminal no...

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Inner Membrane Protein YhcB Interacts with RodZ Involved in Cell Shape Maintenance in Escherichia coli

Cited by 15 publications

References 34 publications

The small RNA, SdsR, acts as a novel type of toxin inEscherichia coli

The small RNA, SdsR, acts as a novel type of toxin inEscherichia coli

Coordinate regulation of the expression of SdsR toxin and its downstream pphA gene by RyeA antitoxin in Escherichia coli

Characterization of inner membrane protein YciB in Escherichia coli: YciB interacts with cell elongation and division proteins

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