Genes 55, alpha gt, 47 and 46 of bacteriophage T4: the genomic organization as deduced by sequence analysis.

The ogr gene product of bacteriophage P2 is a positive regulatory factor required for P2 late-gene transcription. We have determined the nucleotide sequence of the ogr gene, which encodes a basic polypeptide of 72 amino acids. P2 growth is blocked by a host mutation, rpoA09, in the a subunit of DNA-dependent RNA polymerase. The ogr52 mutation, which allows P2 to grow in an rpoAl09 strain, was shown to be a single nucleotide change, in the codon for residue 42, that changes tyrosine to cysteine. The predicted amino acid sequence of the Ogr protein does not show similarity to DNAbinding proteins that are known to affect promoter recognition, to a factors, or to other characterized transcriptional regulatory proteins. We have inserted the ogr gene into a plasmid under control of the leftward promoter and operator of bacteriophage X. Thermal induction of ogr gene expression in this plasmid results in overproduction of a small protein that has been shown by complementation to possess Ogr function.The late genes of the temperate coliphage P2 are organized into four transcription units (1,2) and are regulated in a manner quite different from that of the lambdoid phages. During normal infection, late-gene expression requires the P2 DNA replication genes A and B (2-4) and the host RNA polymerase (5). However, transcription of the P2 late genes initiates at sites that differ in nucleotide sequence from those normally recognized by Escherichia coli RNA polymerase (6, 7). The product of the P2 ogr gene is believed to be a positive regulatory factor required for late-gene expression. Mutations in the ogr gene (8) define a trans-dominant factor that overcomes the block to P2 late-gene transcription imposed by the host rpoAJ09 mutation, which causes a leucine-*histidine substitution in the a subunit of DNA-dependent RNA polymerase (9). Although no amber mutations in the ogr gene have been isolated, it is presumed to be essential by analogy to the B gene of the P2-related phage 186. The existence of P2/186 hybrid phages in which expression ofthe P2 late genes is under 186 B-gene control (10) demonstrates the functional equivalence of the B and ogr gene products. Two amber mutations define the B gene as an essential function needed for 186 gene expression (11), and they map at a site corresponding to that of the ogr mutations in P2. To gain insight into the mechanisms underlying positive control of P2 lategene transcription, we have determined the nucleotide sequence of the P2 ogr gene, inserted the gene into an overproduction vector, and identified the ogr gene product. METHODSPhage and Bacterial Strains. Most strains used for growth of P2 and plasmids were derivatives of E. coli C (12). C-la is prototrophic (13) and C-1055 is used as a nonsuppressing indicator (14). C-2111 and C-2121 (8) carry the rpoA109 mutation. E. coli K-12 strain RR1 (15) carrying the plasmid pRK248cIts (16) was used as the host for derivatives of the plasmid pRC23 (17). P2 virl carries a point mutation that blocks lysogeny (18,19). P2 vir22 has a...

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“…Again, no homologies were found either to the E. coli Ca factors (30,31) or to those encoded by phages SPO1 (32,33), T4 (34), or 429 (35 …”

Section: Resultsmentioning

confidence: 94%

Regulation of bacteriophage P2 late-gene expression: the ogr gene.

Christie¹,

Haggård‐Ljungquist²,

Feiwell³

et al. 1986

Proc. Natl. Acad. Sci. U.S.A.

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“…1) is expected to accentuate the defective DNA replication in the rhoO26 hosts (Fig. 2 (3,14,16,21,63), and in these cases there is a palindromic sequence with an interspersed CUU CGG sequence between the promoter and the structural gene of interest (Fig. 4).…”

Section: Discussionmentioning

confidence: 99%

“…It begins at specific primary or tertiary origins of replication (for reviews, see references 26, 27, 29, 43, and 72). For at least one primary origin, oriA, at 16.5 kilobases on the T4 map, it has been shown that E. coli RNA polymerase initiates primers for leading-strand DNA synthesis (37,49; P. Macdonald We have therefore investigated T4 DNA synthesis and early T4 protein synthesis in rhoO26 mutant bacteria (which are of the NusD phenotype [10]). We observed delayed and reduced synthesis of at least five T4 proteins in T4+-infected rhoO26 cells.…”

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confidence: 99%

Impaired expression of certain prereplicative bacteriophage T4 genes explains impaired T4 DNA synthesis in Escherichia coli rho (nusD) mutants

Stitt

Mosig

1989

J Bacteriol

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The Escherichia coli rhoO26 mutation that alters the transcription termination protein Rho prevents growth of wild-type bacteriophage T4. Among the consequences of this mutation are delayed and reduced T4 DNA replication. We show that these defects can be explained by defective synthesis of certain T4 replicationrecombination proteins. Expression of T4 gene 41 (DNA helicase/primase) is drastically reduced, and expression of T4 genes 43 (DNA polymerase), 30 (DNA ligase), 46 (recombination nuclease), and probably 44 (DNA polymerase-associated ATPase) is reduced to a lesser extent. The compensating T4 mutation goFI partially restores the synthesis of these proteins and, concomitantly, the synthesis of T4 DNA in the E. coli rho mutant. From analyzing DNA synthesis in wild-type and various multiply mutant T4 strains, we infer that defective or reduced synthesis of these proteins in rhoO26-infected cells has several major effects on DNA replication. It impairs lagging-strand synthesis during the primary mode of DNA replication; it delays and depresses recombination-dependent (secondary mode) initiation; and it inhibits the use of tertiary origins. All three T4 genes whose expression is reduced in rhoO26 cells and whose upstream sequences are known have a palindrome containing a CUUCGG sequence between the promoter(s) and ribsome-binding site. We speculate that these palindromes might be important for factor-dependent transcription termination-antitermination during normal T4 development. Our results are consistent with previous proposals that the altered Rho factor of rhoO26 may cause excessive termination because the transcription complex does not interact normally with a T4 antiterminator encoded by the wild-type goF gene and that the T4 goFI mutation restores this interaction.Transcription termination factor Rho plays an important role in regulating gene expression in Escherichia coli and its bacteriophages (for reviews, see references 7, 13, 52, and 69). Some E. coli mutants with defective Rho factors show reduced transcription termination at Rho-dependent termination sites and thus suppress transcriptional polar effects. These mutants can also partly overcome the effects of defective lambda gene N antiterminator function at the lambda Rho-dependent termination sites. In contrast, another class of rho mutants, sometimes called nusD, has a different phenotype.' These mutants do not support the normal growth of wild-type bacteriophage lambda or T4, but their defects can be partially suppressed by mutations in phage antiterminator genes: N in lambda and probably goF, comC-ot, or go-9H (which are probably the same genes [23,64,66]) in T4. It is thought that in these rho mutants, the altered protein is unable to respond to phage-encoded antitermination functions (10, 55, 64).The T4 goF gene is dispensable in many hosts; it maps in a nonessential part of the T4 genome (64, 66). Presumably, in wild-type hosts T4 is far less dependent than phage lambda on antitermination functions because many of the T4 delayed-early gene...

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“…It has been reinserted on the basis of this analysis. It could function as the membrane anchor for the gp46/47 nuclease earlier reported to be a membrane protein (345,731).…”

Section: Hypothetical Proteins With Predicted Cell Membrane Associationsmentioning

confidence: 98%

Bacteriophage T4 Genome

Miller

Kutter

Mosig

et al. 2003

Microbiol Mol Biol Rev

690

801

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SUMMARY Phage T4 has provided countless contributions to the paradigms of genetics and biochemistry. Its complete genome sequence of 168,903 bp encodes about 300 gene products. T4 biology and its genomic sequence provide the best-understood model for modern functional genomics and proteomics. Variations on gene expression, including overlapping genes, internal translation initiation, spliced genes, translational bypassing, and RNA processing, alert us to the caveats of purely computational methods. The T4 transcriptional pattern reflects its dependence on the host RNA polymerase and the use of phage-encoded proteins that sequentially modify RNA polymerase; transcriptional activator proteins, a phage sigma factor, anti-sigma, and sigma decoy proteins also act to specify early, middle, and late promoter recognition. Posttranscriptional controls by T4 provide excellent systems for the study of RNA-dependent processes, particularly at the structural level. The redundancy of DNA replication and recombination systems of T4 reveals how phage and other genomes are stably replicated and repaired in different environments, providing insight into genome evolution and adaptations to new hosts and growth environments. Moreover, genomic sequence analysis has provided new insights into tail fiber variation, lysis, gene duplications, and membrane localization of proteins, while high-resolution structural determination of the “cell-puncturing device,” combined with the three-dimensional image reconstruction of the baseplate, has revealed the mechanism of penetration during infection. Despite these advances, nearly 130 potential T4 genes remain uncharacterized. Current phage-sequencing initiatives are now revealing the similarities and differences among members of the T4 family, including those that infect bacteria other than Escherichia coli. T4 functional genomics will aid in the interpretation of these newly sequenced T4-related genomes and in broadening our understanding of the complex evolution and ecology of phages—the most abundant and among the most ancient biological entities on Earth.

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Genes 55, alpha gt, 47 and 46 of bacteriophage T4: the genomic organization as deduced by sequence analysis.

Cited by 55 publications

References 65 publications

Regulation of bacteriophage P2 late-gene expression: the ogr gene.

Regulation of bacteriophage P2 late-gene expression: the ogr gene.

Impaired expression of certain prereplicative bacteriophage T4 genes explains impaired T4 DNA synthesis in Escherichia coli rho (nusD) mutants

Bacteriophage T4 Genome

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