Two overlapping genes encoding membrane proteins required for bacteriophage N4 adsorption

Kiino, D. R.; Singer, M; Rothman-Denes, Lucia B.

doi:10.1128/jb.175.21.7081-7085.1993

Cited by 24 publications

(22 citation statements)

References 21 publications

(19 reference statements)

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“…Both ManYZ and PtsG are large proteins that have cellular functions in sugar transport. The large (∼85 kDa) inner membrane protein NfrB of unknown cellular function, as well as NfrC (whose gene is now called wecB and is encoded in the same operon as nfrB ), were shown to be essential for Podoviridae E. coli phage N4 adsorption (Kiino et al ., ). The same work showed that the NfrC/Wec B protein is directly involved in the ECA synthesis pathway (see Supporting Information Fig.…”

Section: Discussionmentioning

confidence: 97%

Genes affecting progression of bacteriophage P22 infection in Salmonella identified by transposon and single gene deletion screens

Bohm

Porwollik

et al. 2018

Molecular Microbiology

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Bacteriophages rely on their hosts for replication, and many host genes critically determine either viral progeny production or host success via phage resistance. A random insertion transposon library of 240,000 mutants in Salmonella enterica serovar Typhimurium was used to monitor effects of individual bacterial gene disruptions on bacteriophage P22 lytic infection. These experiments revealed candidate host genes that alter the timing of phage P22 propagation. Using a False Discovery Rate of < 0.1, mutations in 235 host genes either blocked or delayed progression of P22 lytic infection, including many genes for which this role was previously unknown. Mutations in 77 genes reduced the survival time of host DNA after infection, including mutations in genes for enterobacterial common antigen (ECA) synthesis and osmoregulated periplasmic glucan (OPG). We also screened over 2000 Salmonella single gene deletion mutants to identify genes that impacted either plaque formation or culture growth rates. The gene encoding the periplasmic membrane protein YajC was newly found to be essential for P22 infection. Targeted mutagenesis of yajC shows that an essentially full-length protein is required for function, and potassium efflux measurements demonstrated that YajC is critical for phage DNA ejection across the cytoplasmic membrane.

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Section: Discussionmentioning

confidence: 97%

Genes affecting progression of bacteriophage P22 infection in Salmonella identified by transposon and single gene deletion screens

Bohm

Porwollik

et al. 2018

Molecular Microbiology

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“…Since many porins (OmpA, OmpF, OmpC, FhuA, and LamB) have been identified as bacteriophage receptors [5][6][7][8][20][21][22]29], and these have homologous structures [31], we can predict that analogous regions within these Omps might be globally important for phage infection. However, there are few published studies that delineate molecular mechanisms governing phage attachment to these receptors and none to date involving a member of the P22-like phages.…”

Section: Introductionmentioning

confidence: 99%

“…For example, many archaeal, eukaryotic, and bacterial viruses require proteinaceous receptors on the host surface used for attachment [2][3][4][5][6][7][8][9][10][11][12]. In addition, many of these viruses demonstrate plasticity in their binding mechanisms [3,13].…”

Section: Introductionmentioning

confidence: 99%

Key Residues of S. flexneri OmpA Mediate Infection by Bacteriophage Sf6

Porcek

Parent

2015

Journal of Molecular Biology

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Many viruses, including bacteriophage, have the inherent ability to utilize several types of proteinaceous receptors as an attachment mechanism to infect cells, yet the molecular mechanisms that drive receptor binding have not been elucidated. Using bacteriophage Sf6 and its host, Shigella flexneri, we investigated how Sf6 utilizes outer membrane protein A (OmpA) for infection. Specifically, we identified that surface loops of OmpA mediate Shigella infection. We further characterized which residues in the surface loops are responsible for Sf6 binding and productive infection using a combination of in vivo and in vitro approaches including site-directed mutagenesis, phage plaque assays, circular dichroism spectroscopy, and in vitro genome ejection assays. Our data indicate that Sf6 can productively interact with other bacterial OmpAs as long as they share homology in loops 2 and 4, suggesting that these loops may determine host specificity. Our data provide a model in which Sf6 interacts with OmpA using the surface of the protein and new insights into viral attachment through binding to membrane protein receptors.

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“…Approximately 60% of the mutants map to nfrA and nfrB, two tightly linked loci at 12 min on the E. coli linkage map (2) whose coding regions overlap (21). These genes have been previously characterized as an outer membrane protein (NfrA) which presumably is the structural receptor for N4 and an inner membrane protein (NfrB).…”

mentioning

confidence: 99%

A cytoplasmic protein, NfrC, is required for bacteriophage N4 adsorption

et al. 1993

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At least four genes are required for irreversible adsorption of bacteriophage N4. nfrA and nfrB have been characterized previously and encode an outer membrane protein and inner membrane protein, respectively. The nfrC gene product is characterized in detail in this study. We have mapped the nfrD locus to min 52 on the Escherichia coli linkage map. Maxicell analysis of nfrC and a null allele (nfrC2) cloned into a high-copy-number plasmid shows its gene product to be 42 kDa in size. We determined the nfrC nucleotide sequence which predicts a gene product of 42 kDa. Western blots (immunoblots) of Escherichia coli proteins after cellular fractionation show NfrC to be a cytoplasmic protein which is required for irreversible bacteriophage N4 adsorption, an event occurring at the cell surface.Bacteriophage N4 is a lytic phage specific for Escherichia coli K-12 (19). It is unique among the DNA-containing bacteriophages because transcription of the early region of the genome is independent of the activity of the host RNA polymerase. Instead, transcription of the early phage genes is carried out by a virion-encapsulated DNA-dependent RNA polymerase (12). Therefore, the first event in N4 infection requires the injection of both its 72-kbp double-stranded DNA genome and the virion RNA polymerase, a large single-subunit enzyme (M,, 320,000). We have been conducting a genetic analysis of N4 adsorption to learn more about E. coli membrane structure and the penetration of large macromolecular species such as the phage genome and RNA polymerase through the outer and inner membranes.Selection for spontaneously occurring mutations which confer N4 resistance has consistently resulted in mutations mapping to four different loci (20), suggesting the involvement of several E. coli gene products and a relatively complex process. Approximately 60% of the mutants map to nfrA and nfrB, two tightly linked loci at 12 min on the E. coli linkage map (2) whose coding regions overlap (21). These genes have been previously characterized as an outer membrane protein (NfrA) which presumably is the structural receptor for N4 and an inner membrane protein (NfrB). The remaining mutations map to two unlinked loci, nfrC at 85 min and nfrD, which, until this study, had not been precisely mapped.In this article, we characterize the nfrC gene product in detail and present its cloning and the determination of its nucleotide sequence. MATERIALS AND METHODSBacterial strains. (29,38).Cloning nfrC. We screened a XD69 (30) library of E. coli DNA (generously provided by S. Gottesman) for potential clones of nfrC by using the approach described previously (20). We plated the library for single plaques on a lawn of strain KW8801 (carrying the nfrC2 mutation) which had been seeded with N4. Clear plaques representing phage potentially complementing the adsorption defect were picked and purified. Thosc that were turbid in the absence of N4 after replating were lysogenized into KW8801 and tested for their ability to complement the nfrC2 mutation and to integrate at ...

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Two overlapping genes encoding membrane proteins required for bacteriophage N4 adsorption

Cited by 24 publications

References 21 publications

Genes affecting progression of bacteriophage P22 infection in Salmonella identified by transposon and single gene deletion screens

Genes affecting progression of bacteriophage P22 infection in Salmonella identified by transposon and single gene deletion screens

Key Residues of S. flexneri OmpA Mediate Infection by Bacteriophage Sf6

A cytoplasmic protein, NfrC, is required for bacteriophage N4 adsorption

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