Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “Bacteriophage phi 6”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 19 recordsLinked to original sources
PubMed1993

RNA structure and heterologous recombination in the double-stranded RNA bacteriophage phi 6.

Bacteriophage phi 6 has a genome of three segments of double-stranded RNA, designated L, M, and S. A 1.2-kbp kanamycin resistance gene was inserted into segment M but was shown to be genetically unstable because of a high recombination rate between segment M and the 3' ends of segments S and L. The high rate of recombination is due to complementary homopolymer tracts bounding the kan gene. Removal of one arm of this potential hairpin stabilizes the insertion. The insertion of a 241- or 427-bp lacZ' gene into segment M leads to a stable Lac+ phage. The insertion of the same genes bounded by complementary homopolymer arms leads to recombinational instability. A stable derivative of this phage was shown to have lost one of the homopolymer arms. Several other conditions foster recombination. The truncation of a genomic segment at the 3' end prevents replication, but such a damaged molecule can be rescued by recombination. Similarly, insertion of the entire 3-kb lacZ gene prevents normal formation of virus, but the viral genes can be rescued by recombination. It appears that conditions leading to the retardation or absence of replication of a particular genomic segment facilitate recombinational rescue.

Bacteriophage T7↗
PubMed1994

Identification of the packaging regions within the genomic RNA segments of bacteriophage phi 6.

Bacteriophage phi 6 has a genome of three segments of double-stranded RNA enclosed in a procapsid composed of four different proteins. The preformed procapsid is capable of packaging plus-strand transcripts of the genomic segments in an in vitro reaction. The packaging-specific sequences on the RNA molecules are located near the 5' ends. In this study we show that the packaging sequences are different for each of the three segments and that they are of about 250 nucleotides in length. Although these sequences are consistent with some secondary structure, there is no clear structural similarity between the packaging regions of the three segments.

Bacteriophage phi 6↗
PubMed1994

Reconstitution of active replicase in procapsids of the segmented dsRNA bacteriophage phi 6.

Bacteriophage phi 6 has a genome of three segments of double-stranded RNA enclosed in a procapsid composed of four different proteins. The preformed procapsid is capable of packaging plus strand transcripts of the genomic segments in an in vitro reaction. The plus strands then serve as templates for in vitro minus strand synthesis. Procapsids that are missing protein P2 are incapable of minus strand synthesis. In this report we show that incubation with a cell extract containing P2 results in particles with normal amounts of attached P2 and with packaging and replicase activity. Particles lacking P7 have reduced replicase activity which can be augmented by incubation with extracts containing P7, but the amount of attached P7 is small.

Bacteriophage phi 6↗
PubMed1994

RNA structural requirements for stability and minus-strand synthesis in the dsRNA bacteriophage phi 6.

Bacteriophage phi 6 has a genome consisting of three segments of double-stranded RNA designated L, M, and S. Each virion contains one of each genomic segment. Empty procapsids can package plus-strand transcripts of the genomic segments if the 5' regions are intact. Minus-strand synthesis takes place if all three segments are packaged and if the 3' end of the segment is intact. The 3' ends of the segments contain four hairpin structures within a region of high sequence conservation. We now show that removal of parts of this region leads to progressive but limited loss of ability to support minus-strand synthesis. The defective 3' ends can be corrected by heterologous recombination with the termini of other segments. Segments that have small deletions in the conserved region and that support apparently normal minus-strand synthesis are highly recombinogenic.

Bacteriophage phi 6↗
PubMed1980

Physical measurements on the lipid-containing bacteriophage phi 6.

Bacteriophage phi 6 has been studied by small-angle X-ray scattering intensity-fluctuation spectroscopy, analytical ultracentrifugation, and spectroscopy. The sedimentation coefficient (S(0)0,w) is 375 S, the diffusion coefficient (D(0)20, w) is 2.66 x 10(-8) cm 2/s. Using the Svedberg equation and and an estimate of the partial specific volume, the Mr is 1.49 +/- 0.32 x 10(8). A simple model which describes phi 6, is a central sphere consisting of RNA and protein of radius 330 A and an outer shell of low electron density 40 A thick. The RNA may form five concentric shells in the region r = 140-290 A.

Bacteriophages↗
PubMed1995

Packaging of multiple copies of reduced-size genomic segments by bacteriophage phi 6.

Bacteriophage phi 6 has a genome of three segments of double-stranded RNA enclosed in a polyhedral procapsid. The preformed procapsid is capable of packaging plus-strand transcripts of the genomic segments in an in vitro reaction. Packaging of individual segments is dependent upon unique packaging sequences of about 300 nucleotides near the 5' ends of the segments. We have prepared segments L, M, and S with internal deletions that decrease their size by as much as sixfold without affecting either their packaging sequences or their 3' ends. Although packaging of genomic segments is normally very precise, with only one of each in a procapsid, these smaller segments are packaged in multiples such that the total number of nucleotides for each segment class approaches that of the normal genomic segment.

Bacteriophages↗
PubMed1992

Dependence of minus-strand synthesis on complete genomic packaging in the double-stranded RNA bacteriophage phi 6.

Bacteriophage phi 6 has a segmented genome consisting of three pieces of double-stranded RNA (dsRNA). The viral procapsid is the structure that packages plus strands, synthesizes the complementary negative strands to form dsRNA, and then transcribes dsRNA to form plus-strand message. The minus-strand synthesis of a particular genomic segment is dependent on prior packaging of the other segments. The 5' end of the plus strand is necessary and sufficient for packaging, while the normal 3' end is necessary for synthesis of the negative strand. We have now investigated the ability of truncated RNA segments which lack the normal 3' end of the molecules to stimulate the synthesis of minus strands of the other segments. Fragments missing the normal 3' ends were able to stimulate the minus-strand synthesis of intact heterologous segments. Minus-strand synthesis of one intact segment could be stimulated by the presence of two truncated nonreplicating segments. The 5' fragments of each single-stranded genomic segment can compete with homologous full-length single-stranded genomic segments in minus-strand synthesis reactions, suggesting that there is a specific binding site in the procapsid for each segment.

Bacteriophages↗
PubMed1992

Heterologous recombination in the double-stranded RNA bacteriophage phi 6.

Bacteriophage phi 6 contains three double-stranded RNA genomic segments. We have constructed a virus with an insertion of a kanamycin resistance gene in genomic RNA segment M. The virus forms small, turbid plaques, and its genome is unstable. Virus from a single plaque contained from about 0.1 to 10% large clear-plaque forms of the virus; these were usually missing the kanamycin resistance gene, and in many cases, the resulting segment M was larger or smaller than its normal size. Sequence analysis of the genomic RNA of the apparent deletions showed that they were formed by recombination events between segment M and either segment S or L. These heterologous recombination events resulted in the loss of the kanamycin resistance gene from segment M and the replacement of the 3' end of segment M with the 3' end of segment S or L. Although the 3' ends of the single-stranded RNA transcripts of the genomic segments appear to have extensive secondary structure, the sequences at the 3' ends are not involved in the specificity of genomic packaging.

Base Sequence↗
PubMed1995

Acquisition of a fourth genomic segment in bacteriophage phi 6, a bacteriophage with a genome of three segments of dsRNA.

Bacteriophage phi 6 has a genome of three segments of double-stranded RNA enclosed in a polyhedral procapsid. Packaging of individual segments is dependent upon unique packaging sequences near the 5' ends of the segments. We have prepared deletions in segments L and M that decrease their size by half. Phages with these deletions can be propagated on host strains carrying plasmids with complementing genes. The deletion segments are present in two copies per virion. Phage carrying a deletion segment can acquire the transcript of the complementing plasmid if the latter has a packaging sequence. If the packaging sequence is homologous to that of the deletion segment, acquisition occurs at high frequency. If it is heterologous, then recombination exchanges the heterologous packaging sequence for a homologous one or it attaches the transcript to one of the other genomic segments.

Bacteriophages↗
PubMed1976

Temporal origin of viral phospholipids of the enveloped bacteriophage phi 6.

The enveloped bacteriophage phi 6 contains a higher relative level of the negatively charged phospholipid phosphatidylglycerol than is found in the membranes of the host bacterium. During infection of Pseudomonas phaseolicola with phi 6, the level of phosphatidylglycerol synthesis increases significantly. The lipid used to form the viral envelope consists almost entirely of cellular phospholipids synthesized before infection and phosphatidylglycerol synthesized after infection. Based on these and previously published results, a speculative model for this viral envelope formation process is presented.

Bacteriophages↗
PubMed1994

Isolation and characterization of nonsense mutations in gene 10 of bacteriophage phi 6.

Nonsense mutants of bacteriophage phi 6 were isolated by a procedure that involved directed mutagenesis of a cDNA copy of genomic segment M, transcription of this segment, in vitro packaging into procapsids, and transfection of spheroplasts to form viable mutant phage. Recombinant phi 6 viruses that contained amber mutations in two open reading frames, ORF 10 and ORF D, of genomic segment M were isolated. We show that phi 6 protein P10 is the gene product of ORF 10. Further characterization of the phi 6 ORF 10(Am) mutant revealed that phi 6 membrane-associated protein P10 is not required to make enveloped phage particles in infected cells. Enveloped phage particles isolated from a phi 6 ORF 10(Am) infection contained extremely low levels of phi 6 membrane-associated proteins P6 and P3. The low abundance is due to the very low level of P6 synthesis in phi 6 ORF 10(Am)-infected cells. The results suggest that P10 might play a role in regulating the translation of gene 6. Protein P10 was found to be required for host lysis.

Bacteriophage phi 6↗
PubMed1994

Plasmid-directed assembly of the lipid-containing membrane of bacteriophage phi 6.

The nucleocapsid of bacteriophage phi 6 is enveloped within a lipid-containing membrane. The membrane is composed of proteins P3, P6, P9, P10, and P13 and phospholipids. The relationship between membrane protein P9 and morphogenetic protein P12 was studied in the absence of phage infection. cDNA copies of genes 9 and 12 were expressed on plasmids in Pseudomonas syringae pv. phaseolicola. Immunoblotting demonstrated the presence of protein P9 in strains carrying both gene 9 and gene 12 but not in strains with gene 9 alone. In the absence of P12, P9 was found to be unstable. Simultaneous synthesis of proteins P9 and P12 led to the formation of a low-density P9 particle having a buoyant density similar to that of precursor structures composed of phospholipid and proteins isolated from phi 6-infected cells. These results are consistent with results of previous genetic experiments suggesting that P9 and P12 are necessary and sufficient for the formation of the phi 6 envelope. Extensions of P9 at the C terminus do not impair particle formation; however, N-terminal extensions or C-terminal deletions that extend into the hydrophobic region of P9 do impair particle formation.

Amino Acid Sequence↗
PubMed1991

In vitro packaging of the bacteriophage phi 6 ssRNA genomic precursors.

Bacteriophage phi 6 contains three segments of double-stranded RNA within a nucleocapsid. Plasmids containing cDNA copies of the large genomic segment direct the synthesis of viral proteins that assemble into procapsids in Escherichia coli or Pseudomonas phaseolicola. These structures are dodecahedral assemblages of proteins P1, P2, P4, and P7. We report in this paper that these particles are capable of packaging viral single-stranded plus-sense RNA in vitro. The packaging reaction requires the presence of ATP or dATP. Synthesis of minus strands takes place within this filled procapsid in the presence of all four nucleoside triphosphates. Packaged ssRNA is found to be protected from added ribonuclease.

Adenosine Triphosphate↗
PubMed1995

In vitro packaging of individual genomic segments of bacteriophage phi 6 RNA: serial dependence relationships.

Bacteriophage phi 6 has a genome of three segments of double-stranded RNA enclosed in a procapsid composed of four different proteins. The preformed procapsid is capable of packaging plus-strand transcripts of the genomic segments in an in vitro reaction. The packaging of the three segments shows a strong order of dependence in that segment S packages alone, but segment M requires S and and segment L requires S and M for efficient packaging. Packaging of individual segments is dependent on unique packaging sequences of about 200 nucleotides near the 5' ends of the segments. Deletions that invade these regions destroy packaging competence for the particular segment and for the dependent segments as well. In the presence of 2 mM phosphate and at magnesium ion concentrations above 4 mM, packaging becomes progressively more independent and ultimately nonspecific with respect to phi 6 sequences.

Bacteriophage phi 6↗
PubMed1989

Quantitation of the adsorption and penetration stages of bacteriophage phi 6 infection.

The enveloped dsRNA bacteriophage phi 6 uses the pilus of Pseudomonas syringae as its receptor. It enters the host cell by fusion of the virus envelope with the host outer membrane, followed by penetration of the cytoplasmic membrane by the phage nucleocapsid. In this investigation we quantitated the adsorption and penetration of phi 6wt and a host range mutant, phi 6h 1s, to five bacterial strains. Adsorption rate constants were measured for the different phage-host combinations, the constant for phi 6wt with the standard host was 3.3 X 10(10) ml/min. Infections with 14C-labeled phage at different phage/cell ratios were used to measure the numbers of adsorbing and entering virions/sensitive cell. At high phage/cell ratios (200-250) the standard host adsorbed on the average 35-40 wild-type virions/cell, the saturation level being somewhat higher. It was shown that at phage/host cell ratios of 0.1-1 practically every virion produces an infectious center. The average number of entering phage particles per infectious center reached saturation around the phage/cell ratio of 50 and did not exceed 3 for the standard host. The phi 6 preparations used in this study had a specific infectivity of 0.7-0.9.

Adsorption↗
PubMed1975

Intermediates in the biosynthesis of double-stranded ribonucleic acids of bacteriophage phi 6.

Pseudomonas phaseolicola infected with bacteriophage phi 6 synthesized all three viral double-stranded RNA segments, three single-stranded RNAs, and three replicative intermediate-like RNAs in the presence of rifampin. The single-stranded RNA intermediates sedimented and electrophoresed along with melted viral double-stranded RNA, annealed with melted viral double-stranded RNA, and were transient in nature. The relative amounts of the single-stranded RNA intermediates varied during the infection cycle and were altered in the presence of chloramphenicol. The replicative intermediate-like RNAs sedimented faster than double-stranded RNA, failed to enter 2.5% polyacrylamide gels, eluted with double-stranded RNA from a CF-11 cellulose column, were precipitated with single-stranded RNA in 2 M LiC1, and yielded three genome-size pieces of double-stranded RNA upon digestion with RNase. These results are consistent with the hypothesis that complementary strands of the phi 6 double-stranded RNAs are synthesized asynchronously during the infection cycle.

Bacteriophages↗
PubMed1985

Function of pili in bacteriophage phi 6 penetration.

The genome of bacteriophage phi 6, which has a lipid protein envelope, consists of three pieces of dsRNA. Virus infection is initiated by attachment to a phi 6-specific host pilus followed by fusion of the phage membrane and the bacterial outer membrane. In this study we analysed several different phi 6 hosts as well as more than 200 independently isolated phi 6-resistant variants derived from Pseudomonas syringae pv. phaseolicola. It is shown that phi 6-specific pili are coded by genes located in the host chromosome. It appears that pilus reaction is needed to pull the pilus-associated virus through the extracellular polysaccharide of the host and thus to bring it into contact with the outer membrane where membrane fusion can take place.

Adsorption↗