Structure of cryptic λ prophages

Redfield, Rosemary J.; Campbell, Allan

doi:10.1016/0022-2836(87)90289-0

Cited by 18 publications

(12 citation statements)

References 35 publications

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“…Strain BC200 was thought to be deleted for a defective prophage, but map analysis showed it to carry instead an inverted duplication of 45 kb. Strain BC200 was selected as a UV-resistant survivor after a high dose of UV (2), and its duplication may be similar to those seen in cryptic lysogens of E. coli, which arise when lambda lysogens are given high doses of UV (38). We do not know whether the putative prophage genome has been interrupted or deleted by this duplication or whether the lack of prophage expression results from undetectable changes elsewhere in the genome.…”

Section: Discussionmentioning

confidence: 99%

Organization of the Haemophilus influenzae Rd genome

1989

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We present the first complete map of the Haemophilus influenzae genome, consisting of a detailed restriction map with a number of genetic loci. All of the ApaI, SmaI, and RsrII restriction sites (total of 45 sites) were mapped by Southern blot hybridization analysis of fragments separated by pulsed-field gel electrophoresis. Cloned genes were placed on the restriction map by Southern hybridization, and antibiotic resistance loci were also located by transformation with purified restriction fragments. The attachment site of the HP1 prophage was mapped. In addition, the number, locations, and orientations of the six rRNA operons in the H. influenzae chromosome were determined. The positions of conserved restriction sites in these rrn operons confirm that the direction of transcription is 16S to 23S, as in most other bacteria. The widely used strain BC200 appears to contain an unexpected 45-kilobase duplication.A major impediment to genetic analysis of the bacterium Haemophilus influenzae Rd has been the lack of a good genetic map. Although several partial genetic maps have been constructed by using cotransformation frequencies, they include only a small number of markers (about eight antibiotic resistances, an equal number of auxotrophic markers, and a few loci involved in recombination and DNA repair), and the maps are not without discrepancies (19,34,45). Progress in mapping has been impaired for two principal reasons. The first problem is the lack of genetic markers suitable for mapping. H. influenzae is a fastidious microorganism with many growth requirements; as a result, nutritional mutants cannot easily be isolated and characterized. The second problem is that conjugational and transductional mapping, so useful for Escherichia coli and Salmonella typhimurium genetics, has not been well developed for H. influenzae. spp. The natural transformation system of H. influenzae is efficient for mapping closely linked markers, but it is not suitable for large-scale mapping, and cotransformation frequencies depend on the fragment size of the DNA preparation used. Therefore, although a number of genes from H. influenzae have been identified and some have been cloned (see Table 3 for a partial list), it has not been possible to develop ideas about the overall genetic organization of this bacterium.However, within the last few years the development of pulsed-field gel electrophoresis (42) has allowed very large DNA fragments to be separated and physical maps of several bacterial genomes to be constructed (4,12,44,50). Here we present a detailed physical map of the H. influenzae Rd genome, constructed by Southern blot analysis of restriction fragments separated in pulsed-field gels. MATERIALS AND METHODSStrains and culture conditions. The H. influenzae strains used are listed in Table 1. Cells were grown in brain heart infusion broth (Difco Laboratories, Detroit, Mich.) supplemented with hemin (10 ,ug/ml) and NAD (2 ,ug/ml) (sBHI) with gentle shaking at 37°C or on sBHI plates containing 1.5% agar. DNA preparation and rest...

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Section: Discussionmentioning

confidence: 99%

Organization of the Haemophilus influenzae Rd genome

1989

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“…The hybridization data presented in Fig. 3B demonstrate that the mutations resulted from the substitution of the QSR-cos region of phage A by the functional QSR'-cos region from Qsr' cryptic phage (11,24,37 [8] and Becker and Murialdo [2]). We infected wild-type and mutant hosts with the lysis-defective variant, AcI857Sam7, and monitored this cleavage reaction (Fig.…”

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confidence: 99%

HU and integration host factor function as auxiliary proteins in cleavage of phage lambda cohesive ends by terminase

1991

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HU and iategrtion host factor (IHF) are small, basic heterodimeric DNA-binding proteins which participate in transcripion Intatin, DNA replication, and recom . We constructed isogenic Escherichia coli stains in which fIU, THF, or both proteins were absent. Bacteriophage X did not grow in hosts lacing both HU and THE. Phage DNA replication and late gene transcription were normal in the double mutants, but packaging of A DNA was defective. Mature phage DNA molecules were absent, indicating that terminase was unable to linearize K DNA. Phage variants carrying a small substitution near cos or the ohml mutation in the terminase gene, Nul, formed plaques on HU-IHF-strains. We propose that HU or THU is required to establish the higher-order DNA-protein structure at cos that is the substrate for K terminase.DNA-protein complexes with a high degree of organization form at the initiation of DNA replication and are intermediates in several DNA recombination reactions. The ordered structures arise from the concerted assembly of a number of proteins on supercoiled DNA and bring nucleotide sequences located at a distance or on separate molecules into close proximity. In some of these reactions, the DNA bending required to form the complexes is facilitated by low-molecular-weight basic DNA-binding proteins (reviewed by Travers [50]).The best-studied Escherichia coli proteins of this type are the integration host factor (IHF) and HU (reviewed by Drlica and Rouviere-Yaniv [7] and Friedman [14]). IHF is composed of two nonidentical subunits, a and ,B, encoded by the himA (32) and hip (12) genes, respectively. IHF binds in the DNA minor groove (54) and imparts a bend to the DNA in excess of 1400 (27,36,38,49). The binding of IHF to specific DNA sequences at the K attachment site promotes A integration and excision. In a role that is probably related to DNA bending, IHF bound at K promoters can inhibit (22) (10). Although k packaging can take place in THF-hosts, an absolute requirement for IHF can be seen when the phage carries a cos site mutation (cosl54 or cos59) (1,31) or when the host canries a DNA gyrase mutation (15). Phage carrying the ohml mutation in the Nul gene will form plaques on IHF-gyrB hosts (21; also see reference 9).The genes coding for the a and ,B subunits of HU, hupA and hupB, respectively, have recently been cloned (25,26

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“…Therefore, it is likely to be the product of multiple events involving the repeated site-specific integration of prophages, perhaps followed by their stabilization in the host genome by recombination with other resident prophages. Events such as the latter may lead to the stable duplication of genes and may account for the fact that a second copy of the sopE virulence determinant resides within SPI7 (52). Consistent with this idea, SPI7, when it is found in isolates of S. enterica serovar Paratyphi C, appears to lack the sopE gene.…”

Section: Discussionmentioning

confidence: 80%

Precise Excision of the Large Pathogenicity Island, SPI7, inSalmonella entericaSerovar Typhi

et al. 2004

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The large pathogenicity island (SPI7) of Salmonella enterica serovar Typhi is a 133,477-bp segment of DNA flanked by two 52-bp direct repeats overlapping the pheU (phenylalanyl-tRNA) gene, contains 151 potential open reading frames, and includes the viaB operon involved in the synthesis of Vi antigen. Some clinical isolates of S. enterica serovar Typhi are missing the entire SPI7, due to its precise excision; these strains have lost the ability to produce Vi antigen, are resistant to phage Vi-II, and invade a human epithelial cell line more rapidly. Excision of SPI7 occurs spontaneously in a clinical isolate of S. enterica serovar Typhi when it is grown in the laboratory, leaves an intact copy of the pheU gene at its novel join point, and results in the same three phenotypic consequences. SPI7 is an unstable genetic element, probably an intermediate in the pathway of lateral transfer of such pathogenicity islands among enteric gram-negative bacteria.Epidemic recurrences of typhoid fever, caused by Salmonella enterica serovar Typhi, remain among the most costly human infections in terms of both morbidity and mortality (44). S. enterica serovar Typhi is transmitted by contaminated water and food and is an exclusively human pathogen. As with many bacterial infections, the treatment of S. enterica serovar Typhi infection has proven difficult due to the recent emergence of multidrug-resistant strains (54).S. enterica serovar Typhi is closely related to S. enterica serovar Typhimurium, which is among the model eubacteria that can be manipulated rapidly and easily by powerful genetic methods, including phage-mediated genetic exchange or generalized transduction, which was discovered in S. enterica serovar Typhimurium (68). S. enterica serovar Typhimurium can be isolated from a variety of mammals, birds, and reptiles. It has a wide host range and causes a lethal systemic infection in mice yet usually results only in a limited gastroenteritis in humans. In contrast, S. enterica serovar Typhi causes a lethal systemic infection in its exclusively human source and host.To determine the genetic basis of this difference in host range, we are testing the hypothesis that the larger differences between the genomes of these two serovars contribute to their different host ranges. Comparison of the genome sequences of S. enterica serovars Typhi and Typhimurium shows that more than 80% of their sequences are more than 95% identical. They differ primarily by blocks of genes unique to each serovar (7,12,17,18,45,48). The largest difference between these genomes is a 133.5-kb region that includes the genes required for the biosynthesis of the capsular antigen, Vi (36). This region is called Salmonella pathogenicity island 7 (SPI7), or the large pathogenicity island (PI), because has many features in common with PIs found in other gram-negative enteric pathogens (22). SPI7 has a GϩC base composition (49%) significantly different from that of the entire S. enterica serovar Typhi genome (52%), it is bounded by direct repeats overlapping a...

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Structure of cryptic λ prophages

Cited by 18 publications

References 35 publications

Organization of the Haemophilus influenzae Rd genome

Organization of the Haemophilus influenzae Rd genome

HU and integration host factor function as auxiliary proteins in cleavage of phage lambda cohesive ends by terminase

Precise Excision of the Large Pathogenicity Island, SPI7, inSalmonella entericaSerovar Typhi

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