CII-dependent activation of the pRE promoter of coliphage lambda fused to the Escherichia coli galK gene (original) (raw)
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Translation initiation of bacteriophage lambda gene cII requires integration host factor
Journal of bacteriology, 1986
Escherichia coli integration host factor (IHF), a DNA-binding protein, positively regulates expression of the lambda cII gene. Purified IHF stimulates cII protein synthesis in vitro, suggesting a direct role for host factor in cII expression. Further evidence for a direct role for IHF was obtained with operon and gene fusions between cII and lacZ or cII and galE. Analysis of these fusions in vivo demonstrated that IHF is essential for the initiation of cII translation. Replacement of the entire cII coding sequence with lacZ yielded a gene fusion which was still IHF dependent. However, a cII-galE fusion carrying a hybrid ribosome binding region expressed galE in IHF mutants. These results indicate that sequences which make cII translation IHF dependent lie between the ribosome binding region and the initiating codon of cII. Failure to translate cII activates a transcription terminator located within cII and results in polar effects on downstream transcription. This polarity is suppre...
Revisited gene regulation in bacteriophage lambda
Current opinion in genetics & development, 2005
The contribution of bacteriophage lambda to gene control research is far from over. A revised model of the lambda genetic switch includes extra cooperativity through octamerization of the cI repressor protein, mediated by long-range DNA looping. Structural analysis reveals remarkably subtle transcriptional activation by cI. The action of cI, activation by cII, and aspects of antitermination by N and Q all confirm the utility and versatility of simple, weak adhesive interactions mediated by nucleic acid tethers. New genetic and quantitative analysis of the lambda gene network is challenging cherished ideas about how complex behaviours emerge from this regulatory system.
FEMS Microbiology Letters, 1996
It was demonstrated previously that a mutation, rpoA341, in the gene encoding the a subunit of Escherichiu coli RNA polymerase prevents lysogenization by bacteriophage h. The rpoA341 allele is known to be responsible for impaired transcription of some positively regulated E. coli chromosomal operons. Here we demonstrate that the inhibition of lysogenization of the rpoA341 mutant is a result of drastically decreased transcription from positively regulated phage promoters. We were unable to detect any transcripts originating from the CII-activated pz, pi and paQ promoters (important for lysogenic development) in, the phage-infected rpoA341 mutant, in contrast to an otherwise isogenic rpoA+ strain. The results are discussed in the light of other reports showing that activation of the pa promoter by CII protein in vitro is decreased only about fivefold when the native a subunit is replaced by truncated a polypeptides.
Transcription of a bacteriophage lambda DNA site blocks growth of Escherichia coli
Journal of Bacteriology, 1990
The rap mutation in Escherichia coli prevents the growth of bacteriophage lambda. Phage mutations that overcome rap inhibition (bar) have been mapped to loci in the pL operon. We cloned and sequenced three mutations in two of these loci: barIa to the left arm of the lambda attachment site (attP) and barII in the ssb (ea10) gene. The mutations represent single base-pair changes within nearly identical 16-base-pair DNA segments. Each mutation disrupts a sequence of dyad symmetry within the segment. Plasmids carrying a bar+ sequence downstream to an active promoter are lethal to rap, but not rap+, bacteria. The bar sequences isolated from the lambda bar mutants are not lethal. We synthesized a minimal lambda barIa+ sequence, 5'-TATATTGATATTTATATCATT, and cloned it downstream to an inducible promoter. When transcribed, this sequence is sufficient to kill a rap strain.
In vivo transcription studies of coliphage 186
Journal of Virology
The temporal apperance of transcripts from the 186 chromosome has been determined by pulse-labeling at different times after prophage induction and hybridization of RNA extracts to cloned restriction fragments of 186. Studies with different mutants and induction in the presence of chloramphenicol suggested a controlled pattern of transcription and led us to propose the existence of a primary control gene analogous to the lambda gene N.
Molecular and General Genetics MGG, 1978
The activation of the int gene by the cII and cIII gene products was studied by analysing int expression following infection of UV-irradiated cells by various phage mutants. Residual expression of int, probably from PL, takes place in the absence of cII/ cIII activation. Activation of the int gene, like that of the cI repressor gene, is poor at low multiplicities of infection. The mutation intC, which allows constitutive int expression in the lysogenic state, partially relieves the requirement for cII and cIII activation. The kinetics of Int synthesis after addition of the inhibitor rifampicin suggest that the activation occurs at the transcriptional level.
Mutations of coliphage λ affecting the expression of replicative functions O and P
Molecular Genetics And Genomics, 1970
A selection technique, using the thermoinducible prophage ACI857Nsus7 Nsus53, has lead to the characterization of a new class of prophage mutations (called r), which prevent host killing upon thermal induction. N-defective r mutants efficiently complement ~i ~Sa or 40 and P mutants, but not the corresponding mutants of ~i ~:. Complementation data suggest that the 2i z: hybrid fails to provide the positive regulatory mechanism dependent on the 2N-gene product, since it cannot activate the Q gene of a XN-defective mutant. Thus, it seems possible that r mutants cannot express genes 0 and P unless the N-gene product is present in the cell. This interpretation is supported by the fact that r mutants are not defective and form plaques when their N-gene is functional. r mutation confer a clear phenotype, and map in the y-CII region. Results of a density gradient analysis suggest that they result from small insertions of DNA. Induced N-defective r prophages appear to be only poorly transcribed on strand H. Complementation t~sts performed in a strain lysogenie for A indicate that the C 17 mutation can suppress a r mutation in a cis position, even in the absence of the N-gene product. These results suggest that the expression of genes 0 and P, in addition to gene Q, is under the positive regulation of the N-gene product.
Virology, 2003
The bacteriophage cII gene codes for a transcriptional activator protein which is a crucial regulator at the stage of the "lysis-versuslysogeny" decision during phage development. The CII protein is highly toxic to the host, Escherichia coli, when overproduced. However, the molecular mechanism of this toxicity is not known. Here we demonstrate that DNA synthesis, but not total RNA synthesis, is strongly inhibited in cII-overexpressing E. coli cells. The toxicity was also observed when the transcriptional stimulator activity of CII was abolished either by a point mutation in the cII gene or by a point mutation, rpoA341, in the gene coding for the RNA polymerase ␣ subunit. Moreover, inhibition of cell growth, caused by both wild-type and mutant CII proteins in either rpoA ϩ or rpoA341 hosts, could be relieved by overexpression of the E. coli dnaB and dnaC genes. In vitro replication of an oriC-based plasmid DNA was somewhat impaired by the presence of the CII, and several CII-resistant E. coli strains contain mutations near dnaC. We conclude that the DNA replication machinery may be a target for the toxic activity of CII.