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At least 307 records · Page 17Linked to original sources

PubMed1999

Heat-inactivation of plasmid-encoded CI857 repressor induces gene expression from Ind- lambda prophage in recombinant Escherichia coli.

We have observed significant cell lysis upon temperature up-shift of recombinant Escherichia coli cultures harboring CI857-repressed lambda-based expression vectors. This event, that becomes evident about 30-40 min after the heat shock, takes place when using the lambda promoter system in Ind- lysogenic strains, but not in others commonly employed for recombinant gene expression. These results strongly suggest that the thermosensitive CI857 repressor, encoded by the expression vector, competes with CI Ind- molecules for binding to the prophage operator region, allowing for expression of lytic genes from the integrated Ind- viral genome upon temperature up-shift. Transcription of viral lytic genes does not include unspecific expression of a reporter sulA::lacZ gene fusion carried in the prophage genome. These results prompt, however, to carefully evaluate the limitations of expression systems based on pL/pR-CI857 in bacterial strains modified through lambda Ind- gene transfer vehicles.

Bacteriolysis↗

PubMed1996

Sensitization of bacteria to danofloxacin by temperate prophages.

Danofloxacin (CP-76,136) is in a class of agents that inhibit DNA gyrase and trigger induction of the SOS response and temperate bacteriophages. Killing studies against the bovine pathogen Pasteurella haemolytica demonstrated that danofloxacin exhibits particularly rapid killing kinetics. Here, lysogenic Escherichia coli bearing lambda is found to be more sensitive to danofloxacin than nonlysogenic E. coli. Danofloxacin exposure also induced a prophage in P. haemolytica. The potency of danofloxacin against lysogens in likely enhanced by this prophage induction.

Animals↗

PubMed1975

Inhibtory effect of colicin E2 on transport systems of Escherichia coli in the presence of the rex gene of lambda prophage.

Purified colicin E(2) was found to cause marked inhibition of the permeation rate of o-nitrophenyl-galactoside (ONPG) in several lambda-lysogenic strains of Escherichia coli in the presence of chloramphenicol to prevent prophage induction. The inhibitory effect of colicin E(2) on transport systems was analyzed with cells of E. coli CP78(lambda). The dose of colicin E(2) for the half-maximum inhibition of the ONPG-permeation rate was about 9 molecules of the colicin per bacterium under the aerobic condition, which corresponded to about 1 killing unit per bacterium. Kinetics of the transport of [(14)C]methylthiogalactoside suggested that colicin E(2) began to inhibit the influx rate of beta-galactosides within a few minutes after the colicin addition, and the maximum inhibition reached more than 80%. Extensive leakage of intracellular potassium ion and inhibition of l-proline transport also occurred at the same time. Acid solubilization of cellular deoxyribonucleic acid by the colicin was apparently delayed to the initiation of the transport inhibition. The extents of the inhibition of beta-galactoside transport and leakage of potassium ion by the colicin were extensive in cells lysogenic for wild lambda phage or lambdaind(-), whereas the extents were slight in the nonlysogenic cells or cells carrying lambdarex(-) prophage. It was concluded that the sensitization of membrane transport systems of E. coli cells to colicin E(2) was achieved by the presence of the rex gene product of lambda phage.

Biological Transport↗

PubMed1983

Prophage Origin of a Virulent Phage Appearing on Fermentations of Lactobacillus casei S-1.

For protection from the abnormal fermentation of Lactobacillus casei S-1 caused by contamination of a virulent phage, phiFSV, the origin of this phage was studied. Morphologies, viral structural proteins, and DNA structures of three independent isolates of phiFSV were compared with those of phiFSW, which is lysogenized in strain S-1. The results showed (i) that the morphology of phiFSV phages is indistinguishable from that of phiFSW and (ii) that all viral structural components found in phiFSW are present in the particles of phiFSV's. In addition, restriction endonuclease analyses of viral DNA showed that the HindIII-digested fragments of phiFSW DNA, the sum of which covered at least 94.7% of this phage genome, were conserved in the phiFSV DNA digests. Results of Southern filter hybridization of the S-1 and prophage-cured cell (C239) DNAs with phiFSV DNA as a probe revealed that C239 had lost most of the phiFSV DNA sequence, whereas S-1 had about one copy of the phiFSV DNA sequence. These results indicate that virulent phage phiFSV is derived from the lysogenized phage phiFSW. Therefore, the appearance of phiFSV can be eliminated by using the prophage-cured derivative of S-1.

Journal Article↗

PubMed1995

Identification of prophage genes expressed in lysogens of the Lactococcus lactis bacteriophage BK5-T.

Bacteriophage BK5-T is a small isometric-headed temperate phage that infects Lactococcus lactis subsp. cremoris. Northern (RNA) analysis of mRNA produced by lysogenic strains containing BK5-T prophage revealed four major BK5-T transcripts that are 0.8, 1.3, 1.8, and 1.8 kb in size and enabled a transcription map of the prophage genome to be prepared. The position and size of each transcript corresponded closely to the position and size of open reading frames predicted from the nucleotide sequence of BK5-T. Analysis of the transcripts suggested that one of them was derived from the gene encoding the BK5-T integrase and another was from the gene encoding the BK5-T homolog of the lambda cI repressor. Computer analysis of the nucleotide sequence upstream of the BK5-T cI homolog predicted the presence of a pair of divergent promoters and three inverted repeat sequences, features characteristic of temperature-phage immunity regions. By analogy with lambda, the three inverted repeat sequences could be binding sites for cI or Cro homologs and the two divergent promoters could initiate transcription through the BK5-T equivalents of cI and cro.

Amino Acid Sequence↗

PubMed1971

Prophage induction in lysogenic Escherichia coli with simple hydroxylamine and hydrazine compounds.

The prophage-inducing capability of hydroxylamine sulfate and 36 of its derivatives, and of hydrazine dihydrochloride and dihydrazine sulfate and 43 of their derivatives, was determined in Escherichia coli W1709 (lambda). Maximal nontoxic concentrations up to 1 mg/ml were tested. Hydroxylamine sulfate was active at 2.5 mug/ml and the following 17 derivatives were active at concentrations ranging up to 500 mug/ml: alpha-naphthylhydroxylamine, N-hydroxy-2-aminofluorene, oxamyl hydroxamic acid, O-carbamoyl hydroxylamine (isohydroxyurea), N-hydroxyurethane, N-methylhydroxylamine HCl, salicylhydroxamic acid, oxalohydroxamic acid, methoxyamine HCl, ethoxyamine HCl, N, N-diethylhydroxylamine oxalate, formaldoxime, formamidoxime, acetohydroxamic acid, acetaldoxime, acetone oxime, and hydroxyguanidine sulfate. Hydrazine dihydrochloride and dihydrazine sulfate were effective inducers at 5.0 and 2.5 mug/ml, respectively, and the following nine derivatives of them were active at concentrations ranging up to 500 mug/ml: phthalic acid hydrazide, phenylhydrazine HCl, p-nitrophenylhydrazine, p-chlorophenylhydrazine HCl, formylhydrazine, carbohydrazide, semicarbazide HCl, 1-methyl-1-phenyl-hydrazine sulfate, and acetic acid hydrazide. Nineteen hydroxylamine and 34 hydrazine derivatives were ineffective as inducers. Application of the prophage-induction system as a tool for detection of responsive hydroxylamino and hydrazino compounds which may be potential toxicological hazards in the environment is discussed.

Coliphages↗

PubMed1984

Persistence of plasmids, cholera toxin genes, and prophage DNA in classical Vibrio cholerae O1.

Plasmid profiles, the location of cholera toxin subunit A genes, and the presence of the defective VcA1 prophage genome in classical Vibrio cholerae isolated from patients in Bangladesh in 1982 were compared with those in older classical strains isolated during the sixth pandemic and with those in selected eltor and nontoxigenic O1 isolates. Classical strains typically had two plasmids (21 and 3 megadaltons), eltor strains typically had no plasmids, and nontoxigenic O1 strains had zero to three plasmids. The old and new isolates of classical V. cholerae had two HindIII chromosomal digest fragments containing cholera toxin subunit A genes, whereas the eltor strains from Eastern countries had one fragment. The eltor strains from areas surrounding the Gulf of Mexico also had two subunit A gene fragments, which were smaller and easily distinguished from the classical pattern. All classical strains had 8 to 10 HindIII fragments containing the defective VcA1 prophage genome; none of the Eastern eltor strains had these genes, and the Gulf Coast eltor strains contained a different array of weakly hybridizing genes. These data suggest that the recent isolates of classical cholera in Bangladesh are closely related to the bacterial strain(s) which caused classical cholera during the sixth pandemic. These data do not support hypotheses that either the eltor or the nontoxigenic O1 strains are precursors of the new classical strains.

Bacteriophages↗

PubMed1998

A globally disseminated M1 subclone of group A streptococci differs from other subclones by 70 kilobases of prophage DNA and capacity for high-frequency intracellular invasion.

The M1inv+ subclone of M1 group A streptococci that spread globally in the late 1980s and early 1990s was previously identified by restriction fragment length polymorphism (RFLP), M protein, and SpeA exotoxin sequence analyses. Strains representing this subclone were characterized with regard to carriage of bacteriophage and capacity to invade cultured human epithelial cells. The M1inv+ subclone was found to harbor two entirely different prophages, phage T13 and phage T14, which together supplement its genome with nearly 70 kb of DNA. Phage T14 encodes the SpeA exotoxin and is closely related to the classic converting phage T12. Plaque-forming characteristics and RFLP analyses of phages T13 and T14 were compared to each other and to phage T12. Other subclones of M1, isolated in the 1970s to the early 1980s, lacked both prophages. The M1inv+ subclone was previously reported to be efficiently internalized by human epithelial cells. This potential was confirmed and expanded by comparing a variety of clinical isolates. The capacity for high-frequency invasion of epithelial cells was not transmitted to a laboratory strain of group A streptococci by the above-mentioned bacteriophages.

Clone Cells↗

PubMed2001

Mu-like Prophage in serogroup B Neisseria meningitidis coding for surface-exposed antigens.

Sequence analysis of the genome of Neisseria meningititdis serogroup B revealed the presence of an approximately 35-kb region inserted within a putative gene coding for an ABC-type transporter. The region contains 46 open reading frames, 29 of which are colinear and homologous to the genes of Escherichia coli Mu phage. Two prophages with similar organizations were also found in serogroup A meningococcus, and one was found in Haemophilus influenzae. Early and late phage functions are well preserved in this family of Mu-like prophages. Several regions of atypical nucleotide content were identified. These likely represent genes acquired by horizontal transfer. Three of the acquired genes are shown to code for surface-associated antigens, and the encoded proteins are able to induce bactericidal antibodies.

ATP-Binding Cassette Transporters↗

PubMed1972

Cell division and prophage induction in Escherichia coli: effects of pantoyl lactone and various furan derivatives.

Strain T-44 is a thermosensitive mutant of Escherichia coli in which both cell division and prophage repression are altered at elevated temperatures. The effects of various ribosides, pantoyl lactone, and the furfural derivatives nitrofurazone and 5-methyl furfural suggest that some low-molecular-weight compound is important in the control of cell division and prophage repression in this strain. This low-molecular-weight compound may have a five-membered oxygen-containing ring as part of its structure.

Cell Division↗

PubMed1978

Chromosome replication in an Escherichia coli dnaA mutant integratively suppressed by prophage P2.

Escherichia coli CRT4624-P2sig5 is a dnaA mutant in which integration of the prophage P2sig5 has occurred at the attP2II site (min 85). This strain was integratively suppressed, and when cells were shifted to 42 degrees C replication was initiated at a site in or near the P2 prophage. Initially, this replication occurred primarily in the direction that corresponds to the clockwise direction on the genetic map. Replication also occurred in the counterclockwise direction, but the initiation of replication in this direction occurred approximately 40 min later than the initiation of replication in the other direction. Because of this delay, the replication forks that traveled in the clockwise direction were the first to arrive in the region of the replication terminus. These replication forks ceased replication near the aroD locus (min 37), and it is proposed that the replication terminus is between the aroD and rac loci (min 31). A model is proposed for the cycle of chromosome replication in this strain at 42 degrees C.

Chromosomes, Bacterial↗

PubMed1978

Viral R plasmid Rphi6P: properties of the penicillinase plasmid prophage and the supercoiled, circular encapsidated genome.

Properties of the viral R plasmid Rphi6P are described. As a temperate bacteriophage, it plaques on the facultative phototroph Rhodopseudomonas sphaeroides. Under aerobic conditions the phage had a latent period of 180 min, a burst time of 200 min, and a burst size of 15 to 20 particles per infective center. The encapsidated viral genome occurred as a supercoiled, circular DNA duplex with a mean contour length of 16.5 +/- 10 micron. Percent guanine plus cytosine, as calculated from thermal denaturation profiles, was 63.5. Mitomycin C-induced loss of the prophage suggested an extrachromosomal location in the host cell. Use of this curing agent enabled the isolation of a plasmid-free strain of R. sphaeroides. Biophysical analysis of the plasmid-free strain lysogenized with Rphi6P confirmed that the prophage occurred as a plasmid in the host cell.

Adsorption↗

PubMed1985

Mutation-dependent suppression of recB21 recC22 by a region cloned from the Rac prophage of Escherichia coli K-12.

Using pBR322 as a vector, we cloned a 5.95-kilobase fragment of the Rac prophage together with 1.70 kilobases of a flanking Escherichia coli chromosome sequence. The resulting plasmid (pRAC1) was unable to suppress the mitomycin and UV sensitivity and recombination deficiency of a recB21 recC22 strain. Five spontaneous mitomycin-resistant derivatives contained deletion mutant plasmids. These plasmids also suppressed the UV sensitivity and recombination deficiency of their recB21 recC22 hosts. All five deletions were contained within a 2.45-kilobase EcoRI-to-HindIII segment of the plasmid. By substituting the corresponding 2.45-kilobase EcoRI-toHindIII fragments of Rac prophage isolated from sbcA+, sbcA6, and sbcA23 strains for the shortened segment of one of the deletion mutant plasmids, we were able to show that sbcA mutations map in this region. Also in this region is the site (or closely linked sites) at which previous studies had shown that insertion of Tn5 and IS50 leads to suppression of recB21 recC22. The sequence in this region that must be altered or circumvented to allow suppression is discussed. Also presented are data correlating the expression of nuclease activity with the degree of suppression.

Bacteriophage lambda↗

PubMed1990

Characterization of PBSX, a defective prophage of Bacillus subtilis.

PBSX, a defective Bacillus subtilis prophage, maps to the metA-metC region of the chromosome. DNA (33 kilobases) from this region of the chromosome was cloned and analyzed by insertional mutagenesis with the integrating plasmid pWD3. This plasmid had a promoterless alpha-amylase gene (amyL) that provided information on the direction and level of transcription at the site of integration. Transcription under the control of the PBSX repressor proceeded in the direction metA to metC over a distance of at least 18 kilobases. Electrophoretic analysis of proteins produced by different integrant strains upon PBSX induction and by fragments subcloned in Escherichia coli allowed the identification of early and late regions of the prophage. A set of contiguous fragments directing mutagenic integration suggested that the minimum size of an operon that encodes phage structural proteins is 19 kilobases. The adaptation of PBSX transcriptional and replicational functions to a chromosomally based, thermoinducible expression system is discussed.

Bacillus subtilis↗

PubMed1991

Association of a retroelement with a P4-like cryptic prophage (retronphage phi R73) integrated into the selenocystyl tRNA gene of Escherichia coli.

A new multicopy single-stranded DNA (msDNA-Ec73) was found in a clinical strain of Escherichia coli. Retron-Ec73, consisting of an msDNA-coding region and the gene for reverse transcriptase (RT), was found to be a part of a 12.7-kb foreign DNA fragment flanked by 29-bp direct repeats and integrated into the gene for selenocystyl-tRNA (selC) at 82 min on the E. coli chromosome. Except for the 2.4-kb retron region, the integrated DNA fragment showed remarkable homology to most of the bacteriophage P4 genome. Among the phage genes found in this element, however, the integrase gene had very low identity (40%) to P4 integrase, indicating that the cryptic prophage associated with the retroelement has its own unique site-specific integrase different from P4 integrase. Recently, we have shown that P2 phage can act as a helper to excise the cryptic prophage and to package its genome into an infectious virion. The newly formed phage (retronphage phi R73) can also lysogenize a new host strain, reintegrating its genome into the selC gene and enabling the newly formed lysogen to produce msDNA-Ec73 (S. Inouye, M. G. Sunshine, E. W. Six, and M. Inouye, Science 252:969-971, 1991).

Amino Acid Sequence↗

PubMed1993

Genetic and molecular analyses of the C-terminal region of the recE gene from the Rac prophage of Escherichia coli K-12 reveal the recT gene.

The nucleotide sequence of the C-terminal region of the recE gene of the Rac prophage of Escherichia coli K-12 reveals the presence of a partially overlapping reading frame we call recT. Deletion mutations show that recT is required for the RecE pathway of conjugational recombination. By cloning recT with a plasmid vector compatible with pBR322, we showed by cis-trans tests that the portion of the recE gene encoding ExoVIII DNA nuclease activity is also required for RecE pathway conjugational recombination. The recT gene can replace the redB gene of lambda for recA-independent plasmid recombination. A Tn10 insertion mutation previously thought to be in recE is located in recT and is renamed recT101::Tn10. Discrepancies between the molecular mass estimates of wild-type ExoVIII protein determined from mobility in sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and calculated from the predicted amino acid sequence are discussed. The hypothesis that wild-type ExoVIII protein results from fusion of RecE and RecT proteins is disproved genetically, thus supporting a previous hypothesis that the discrepancies are due to abnormal protein mobility in SDS-PAGE. A computer-performed scan of the bacteriophage nucleotide sequence data base of GenBank revealed substantial similarity between most of recE and a 2.5-kb portion of the b2 region of lambda. This suggests interesting speculations concerning the evolutionary relationship of lambda and Rac prophages.

Amino Acid Sequence↗

PubMed1998

RecA-independent pathways of lambdoid prophage induction in Escherichia coli.

Two Escherichia coli genes, expressed from multicopy plasmids, are shown to cause partial induction of prophage lambda in recA mutant lysogens. One is rcsA, which specifies a positive transcriptional regulator of the cps genes, which are involved in capsular polysaccharide synthesis. The other is dsrA, which specifies an 85-nucleotide RNA that relieves repression of the rcsA gene by histone-like protein H-NS. Genetic contexts known to increase Cps expression also cause RecA-independent lambda induction: the rcsC137 mutation, which causes constitutive Cps expression, and the lon and rcsA3 mutations, which stabilize RcsA. Lambdoid phages 21, phi80, and 434 are also induced by RcsA and DsrA overexpression in recA lysogens. Excess lambda cI repressor specifically blocks lambda induction, suggesting that induction involves repressor inactivation rather than repressor bypass. RcsA-mediated induction requires RcsB, the known effector of the cps operon, whereas DsrA-mediated induction is RcsB independent in stationary phase, pointing to the existence of yet another RecA-independent pathway of prophage induction.

Bacterial Proteins↗

PubMed2001

Characterization of VPI pathogenicity island and CTXphi prophage in environmental strains of Vibrio cholerae.

Environmental isolates of Vibrio cholerae of eight randomly amplified polymorphic DNA (RAPD) fingerprint types from Calcutta, India, that were unusual in containing toxin-coregulated pilus or cholera toxin genes but not O1 or O139 antigens of epidemic strains were studied by PCR and sequencing to gain insights into V. cholerae evolution. We found that each isolate contained a variant form of the VPI pathogenicity island. Distinguishing features included (i) four new alleles of tcpF (which encodes secreted virulence protein; its exact function is unknown), 20 to 70% divergent (at the protein level) from each other and canonical tcpF; (ii) a new allele of toxT (virulence regulatory gene), 36% divergent (at the protein level) in its 5' half and nearly identical in its 3' half to canonical toxT; (iii) a new tcpA (pilin) gene; and (iv) four variant forms of a regulatory sequence upstream of toxT. Also found were transpositions of an IS903-related element and function-unknown genes to sites in VPI. Cholera toxin (ctx) genes were found in isolates of two RAPD types, in each case embedded in CTXphi-like prophages. Fragments that are inferred to contain only putative repressor, replication, and integration genes were present in two other RAPD types. New possible prophage repressor and replication genes were also identified. Our results show marked genetic diversity in the virulence-associated gene clusters found in some nonepidemic V. cholerae strains, suggest that some of these genes contribute to fitness in nature, and emphasize the potential importance of interstrain gene exchange in the evolution of this species.

Amino Acid Sequence↗