Post Transcriptional Controls

As with other phages, post-transcriptional mechanisms modulate expression of T4 genes in a variety of ways (i) by modulating translation initiation via potential secondary RNA structures and or by coupling translation of two or more genes, (ii) by translational repression, (iii) by proteins affecting ribosome structure and function, (iv) by processing of primary transcripts, and (v) by the rate of degradation. In addition, expression of at least four T4 genes requires splicing, or in a...

Prophage Insertion by Transposition

The most familiar example of insertion by transposition is coliphage Mu-1. There are fundamental differences between the life cycles of Mu-1 and l. In Mu-1, there is no circular form of the viral DNA. Also, viral DNA is always inserted into host DNA, even in the virion. Both replication and insertion result from transposition of viral DNA from one host site to another. Transposition is diagrammed in figure 7-5. Transposition has two possible outcomes, replicative and nonreplicative, of which...

FX174 et al the Microviridae

BRENTLINGER, APRIL D. BURCH, MIN CHEN, SUSAN HAFENSTEIN, ERICA MOORE, CHRISTOPHER R. NOVAK, AND ASAKO UCHIYAMA Sometime in the 1920s, a plaque was placed in a test tube labeled 174. It must have had a certain luster. Since then that phage has challenged genetic, biochemical, and biophysical dogmas. From the very beginning, it was unique. It was unusually tiny, readily passing through the smallest of ultrafilters. Sertic and Bulgakov (111) used its size to define a race...

Evolution of Phage Latent Period

From an ecological perspective we can divide the members of phage populations into two groups prereproductive and reproductive. Prereproductive phage are those engaged in either adsorption (including the extracellular search for susceptible bacteria) or the eclipse during these periods the phage is not generating mature phage progeny. Reproductive phage, by contrast, are those infecting bacteria during the period of phage-progeny maturation. For phage that must lyse their host bacteria to...

Phage Assembly

Filamentous phage assembly is a secretory process. Assembly occurs in the cytoplasmic membrane, and nascent phages are secreted from the cell as they assemble (figure 12-5). All eight of the phage-encoded proteins that are directly involved in assembly are integral membrane proteins. This includes three nonvirion proteins pI and its restart partner, pXI, in the cytoplasmic membrane (69) and pIV in the outer membrane (12) and the five viral coat proteins, which reside in the cytoplasmic membrane...

Head Structure and Assembly

The head assembly pathway is summarized in figure 18-6. The completed T4 head is composed primarily of the major cleaved capsid protein, gp23*, arranged in an icosahedral surface lattice of T 13, elongated along a 5-fold axis (267). The length of the T4 head is defined by a number, Q, related to the icosahedral T number. An illustration of Q numbers is shown in figure 18-7 together with models of someT4 head lengths. The 5-fold vertices, except for one, are occupied by the cleaved vertex...

Plasmaviridae Shape

AParentheses indicate approximate numbers of poorly characterized isolates that may or may not represent independent species. bArchaea. aParentheses indicate approximate numbers of poorly characterized isolates that may or may not represent independent species. bArchaea. known leviviruses are plasmid-specific coliphages that adsorb to F or sex pili. They have been divided, by serology and other criteria, into two genera. Several as yet unclassified leviviruses are specific for other plasmid...

Structure of the Phage Particle

The image of a filamentous phage obtained by atomic force microscopy is shown in figure 12-2. They are about 6.5 nm in diameter, with a length determined by the size of the genome, normally 6-7 kb of single-stranded DNA. The 6400-nucleotide Ff genome is encapsidated in a 930 nm particle a 221-nucleotide microphage variant is only 50 nm long (152), and the presence of cloned DNA in the phage genome makes the particle proportionately longer. Although there is no theoretical limit to the amount of...

The HK022 Nun Protein

Although not identified as a regulatory protein for its own genome, the action of the HK022 nun gene product is so tied to the action of l N that it warrants discussion in this review. The HK022 nun gene is located in the same relative position on the HK022 genome as the various N genes are located on the genomes of other lambdoid phages. However, the Nun protein does not act as an antitermination protein for HK022 but, instead, acts by terminating transcription on the l genome at sites not...

Prophages Inserted by Reciprocal Site Specific Recombination

L is one of the many phages that insert their DNA into the host chromosome by reciprocal recombination between the chromosome and a circular form of the phage DNA. The overall reaction comprises cointegration of the two circles. The recombination takes place between a unique site on the phage DNA and a unique site on the E. coli Table 7-1 Modes of Prophage Maintenance Insertion By site-specific recombination Coliphage l Salmonella phage P22 Mycobacterial phage L5 Vibrio phage CTX Phi...

DNA Replication

Positive polarity single-stranded DNA replication strategies are complex, occurring in three separate stages. These processes will be briefly summarized a more complete discussion can be found in the previous edition of this book (62). Stage I DNA replication involves the conversion of the single-stranded genome into a covalently closed, double-stranded, circular molecule, called RF I DNA (replicative form one DNA). Purified single-stranded Microviridae DNA produces progeny in transfection...

Lysis Without Lysozyme

It appears that all phages with double-stranded nucleic acid genomes use the holin-endolysin system for lysis. This must reflect a powerful evolutionary pressure for optimization of the lytic event, both in terms of efficiency of release of the progeny virions and, probably more so, in terms of terminating the vegetative cycle at a time appropriate for the particular host and environment. However, no lysozyme is encoded or produced by three classes of very simple lytic phages with small,...

Lysis Lysogeny Decision

As described above, CI must be present to stimulate expression of PRM in the lysogenic state. In a cell that is following the lysogenic pathway, the initial burst of CI is provided by expression of another promoter, PRE (9, 16). Regulation of this promoter involves physiological events that are not fully understood. The direct positive regulator of PRE is the phage protein CII, which binds at PRE (Figure 8-2C) and activates its expression. The level of CII in the cell is in turn under complex...

Harald Brussow And Frank Desiere

Many phage researchers believe that phages are as old as their bacterial hosts. If this hypothesis is true, then we have to postulate elements of vertical evolution for phages. In view of the postulated antiquity of these relationships we might not expect sequence similarities between more distantly related phages. Comparative phage genomics can reveal DNA sequence, protein sequence, or gene map similarities in the absence of sequence similarities between increasingly diverged phages. For even...

Coda

Transcription antitermination can serve two different roles useful to the phage. First, it allows the phage to acquire new genes even if the genes have associated transcription terminators. If an acquired gene locates within an operon, the antitermination mechanism enables transcription to reach downstream genes even if the acquired gene has an associated downstream terminator. For example, in phages carrying genes encoding Shiga toxin, even though the stx genes have an associated promoter,...

Structure of Packaged DNA

Whatever the mechanism of DNA translocation, it must overcome the energetic barrier of compacting the DNA and deliver the DNA into the prohead to confer the proper structure and organization within the head. DNA packaging in dsDNA phages is endothermic. Analogies are often made between DNA packaging and the collapse of DNA into a torus, or DNA toroid, in the presence of polyamines such as spermidine (29) or hexamine cobalt (98). While the final structures share structural similarities, such as...

Perspectives and Directions

In recent years remarkable strides have been made in our understanding of the molecular basis of phage lysis and its regulation. The progress has raised many new questions. That there are at least two general strategies for lysis is clear. All complex phages seem to use holin-endolysin lysis, whereas two of the prototype small ssDNA and ssRNA phages encode proteins which act as inhibitors of cell wall synthesis. The diversity of holins has always been a stunning, and somewhat daunting feature...

Lysis Systems At Least Two Strategies

Most bacteria have a murein cell wall that constitutes the principal barrier to host lysis. Compromising the cell wall is thus the fundamental goal for lytic processes. Two general strategies to accomplish lysis have been described (121). Phages with double-stranded nucleic acid genomes use the holin-endolysin strategy. In this scheme, the phage elaborates a murein-degrading enzyme, an endolysin (also known as lysozyme), specifically dedicated to lysis, and a second, membrane-embedded protein,...

Jan Van Duin Nina Tsareva

The single-stranded RNA coliphages were discovered by Tim Loeb and Norton Zinder in 1961 as the result of a search for phages whose infection cycle depends on E. coli F-pili, normally used for bacterial conjugation. Loeb and Zinder plated filtered samples of raw New York City sewage on E. coli strains and screened for phages that would produce plaques on male (F+) but not female (F ) bacteria. The first isolate, named f1, turned out to be a filamentous phage with a single-stranded DNA genome...

PRD1 Dissecting the Genome Structure and Entry

BUTCHER, JAANA K. H. BAMFORD, AND DENNIS H. BAMFORD There are six extremely similar phage isolates, from different parts of the world, that all infect Gramnegative bacteria harboring a conjugative plasmid. One of these viruses is PRD1. Viruses with this morphology (protein capsid surrounding a membrane vesicle containing the linear double-stranded DNA genome) also infect Grampositive hosts. These viruses have been classified together forming the Tectiviridae family...

Myoviridae

The same pattern of overall DNA sequence alignment and graded relatedness was also observed in phages with a contractile tail (Myoviridae). E. coli phage Mu (representative of the Mu-like genus in the Myoviridae family) and E. coli O157 Sakai prophage Sp18 could be aligned over about half of the genome length at the DNA level in a mosaic fashion (figure 4-4F). Mu shared with the Shewanella prophage MuSo2 protein sequence similarity, but only overall genome organization in the absence of...

The NUT Site

Information gained from genetic and structural studies as well as genome comparisons has extended our understanding of the nature and function of interactions at NUT RNA. The nut sites have been divided into three components boxA, a spacer, and boxB (figure 9-2A). Although the interactions at NUT involve a complex formed between all of the components, the following discussion will focus on each of the component sites individually. A comparison of boxA sites yielded a consensus sequence,...

References

Structure of the lipid-containing bacteriophage PRD1 disruption of wild-type and nonsense mutant phage particles with guanidine hydrochloride. J.Virol. 44 1031-1038. 2. Bamford, D. H., and H.-W. Ackermann. 2000. Family Tectiviridae, pp. 111-116. In M. H. V van Regenmortel, C. M. Fauquet, D. H. L. Bishop, E. B. Carstens, M. K. Estes, S. M. Lemon, J. Maniloff, M. A. Mayo, D. J. McGeoch, C. R. Pringle, R. B.Wickner (eds.) Virus Taxonomy Classification and...

Contributors

Abedon, Department of Microbiology, Ohio State University, Mansfield, OH 44906, USA Hans-W. Ackermann, Department of Medical Biology, Faculty of Medicine, Laval University, Qc G1K7p4, Quebec, Canada Sankar Adhya, Laboratory of Molecular Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892, USA Juan C. Alonso, Departmento de Biotechnologia Microbiana, Centro Nacional de Biotecnologia-Consejo Superior de Investigaciones...

Reduced Phage Productivity

Host density impacts on community stability by affecting the peak phage densities that follow community-wide host lysis. With more phage than hosts within a batch-culture system, eventually all sensitive bacteria may become adsorbed and lysed (10, 29). However, in continuous culture there will be decay in free-phage densities due to outflow from the growth vessel. Since the rate at which host cells are found by free phage is a function of free-phage density (1), there is a race between the...

Antiholin Diversity and Raison dEtre

As stated above, many holin genes have dual-start motifs, with two potential start codons separated by at least one codon for a basic amino acid. In the few other cases tested (P22 13, 21 S, and f 29 14), eliminating codon 1 accelerates the onset of lysis, which is presumptive evidence that the dual start functions analogously to that of S (6, 77, 102). At least for S21, the holin gene of lambdoid phage 21, the longer product has been shown to have antiholin character, based on the N-proximal...

Receptor Recognition

PRD1 belongs to the class of broad-host-range, donor-specific phages, which infect cells only when IncP-, IncN-, or IncW-type multiple drug resistance conjugative plasmids are present. Among the hosts are several opportunistic human pathogens such as E. coli, Salmonella enterica, and Pseudomonas aeruginosa. Plasmid functions in the phage life cycle are dispensable after the genome has entered the cell (23, 43). These plasmids, the best studied of which is the IncP plasmid RP4 (50), encode the...

Contribution of Early Adsorbers to Phage Population Growth

Phage adsorption occurs essentially as an exponential decay in free-phage density (figure 5-2A). For any phage cohort released at a given moment into a population of potential hosts, phage adsorption occurs such that some constant fraction of remaining free phage will adsorb over any given interval. As a consequence, more phage from a given cohort will adsorb during an earlier interval compared with a later interval. Furthermore, if by chance a phage adsorbs a host earlier rather than later,...

Regulation in a Lysogen

In a lysogenic bacterium, the genes of the phage lytic cycle must be turned off, or the cell would die. Turnoff is caused by a phage-coded repressor (usually a protein, but in some cases, as with phage P4, an antisense RNA). The repressor prevents transcriptional initiation from key phage promoters. This turns off almost all phage transcription. Repressor acts directly on only a few promoters (in l, there are two). These are the promoters from which lytic development is initiated. Phage genes...

Hansw Ackermann

Bacteriophages or phages were discovered twice in a short time. The British pathologist Frederick William Twort described in 1915 the glassy transformation of Micrococcus colonies by an unknown agent. Felix Hubert d'Heer-elle, a French Canadian then working at the Pasteur Institute of Paris, observed the destruction of Shigella bacteria in broth (5). Contrary to Twort, he clearly recognized the viral nature of this phenomenon and devoted the rest of his scientific career to it. He coined the...

How Do Holins Work

The foregoing has promoted the idea that holins are remarkable proteins, honed by powerful evolutionary forces to be the simplest and most adjustable of, if not clocks, biological timers. Moreover, it also seems clear that holins are sensitive to the energy state of the membrane and thus, perhaps, constitute the simplest of gated membrane assemblies. The reader has no doubt noticed, however, that there is very little mechanistic insight into how holins work, however elegant and amusing the...

FX174 E

The fX174 genome has only 10 genes, and the lysis gene, E, spans only 91 codons in an alternate reading frame embedded in an essential gene D encoding a capsid scaffolding protein (figure 10-9). E is a membrane protein its first 35 residues define the lytic function and the membrane insertion domain. Although its topology is uncertain, several lines of evidence indicate the C-terminus is cytoplasmic. Replacement of the entire C-terminal cytoplasmic domain by b-galactosidase, GFP, or any folded...

Modification and Restriction of T4 DNA

In the following discussion the term restriction is used in its broadest meaning and is not limited to type II restriction enzymes. The complex modifications and restrictions of T4 DNA and of other DNA by T4 can be best rationalized as a result of ongoing evolution that is accelerated by strong selective forces and exchanges between the genomes of T4 and its host, including plasmid and prophage genomes that reside in the host. As mentioned above, T4 DNA contains HMC instead of C residues. This...

Filamentous Phage Display

Phage display is an extraordinarily powerful method of linking phenotype and genotype in the selection of proteins or peptides (see also chapter 44). The principle is very simple a DNA sequence is inserted into a gene encoding a phage coat protein so as to make a fusion protein in which the coat protein now displays the protein product of the inserted sequence. If a ligand is bound to a solid support, those phages expressing a protein or peptide that binds to the ligand will be immobilized, the...

Gisela Mosig Fred Eiserling

Bacteriophage Diagram

BacteriophagesT2,T4, andT6 were among seven Escherichia coli phages selected by Max Delbr ck to study fundamentals of viral replication in a limited number of model viruses. These studies led to the first formulation of many concepts that are now accepted foundations of molecular biology the fundamental differences between growth of viruses and cells figure 18-1 109 the demonstration that nucleic acids of virus particles suffice to establish infection and to direct synthesis of complete virions...

Prophage Genomics

At the time of this writing, the GenBank phage database comprises 200 complete phage genome sequences. An equivalent number of phage sequences were passively acquired as prophages in bacterial genome sequencing projects. The scientific value of these prophage sequences was only recently recognized 13, 14 . It goes beyond their potential to double the content of the current phage database and to correct a bias of the database towards selected phage systems coliphages, dairy phages,...

Info

Based on the organism list of the National Center for Biotechnology Information NCBI database http www.ncbi.nlm.nih.gov taxonomy 15 and on phage counts from 3 . Total phage numbers are lower than in table 2-1 because rRNA data are unavailable for some phage hosts. CFP, cubic, filamentous, and pleomorphic phages. aIncluding a phage-like particle. so far, without phages. Most phages are found in easily grown and medically or industrially important bacteria 1. Firmicutes with high G-C coryneforms,...

Tls

Figure 17-3 Consensus sequence logos of the T1 and TLS 21 nucleotide direct repeats. Produced using WebLogo Figure 17-3 Consensus sequence logos of the T1 and TLS 21 nucleotide direct repeats. Produced using WebLogo of rho-independent terminators. In coliphage T4 these sites frequently contained a UUCG or GNRA loop sequence 70 that none of the putative T1 TLS terminators possess. The presence of transcriptional terminators led us to speculate that either a transcriptional read-through system...

R1

Figure 15-12 Comparison of RNA secondary structures in the 3' UTR in four representative RNA phages. SP is a species IV allolevivirus. PP7 and AP205 resemble SP in their simple 3' UTR structure. MS2 has a more elaborate folding with many extra elements shaded . The stopcodon of replicase is boxed R stop . Also boxed is a conserved sequence in the top of U1 UGCUU . In Qb and SP these nucleotides form a pseudoknot with their complement 1200 nucleotides upstream figure 15-6 . For MS2, PP7, and...

Gregory J German Rajeev Misra Andrew M Kropinski

Haemophilus Phage Hp2

Although T1 got its initial fame from its use, in 1943, by Salvador Luria and Max Delbr ck in their landmark fluctuation test 64 , an entirely different reason has resulted in its notoriety since then. This phage's resistance to desiccation results in its persistence for weeks in an aerosol which in turn created a real nightmare for laboratories engaged in research employing E. coli K-12 strains. It was not uncommon to hear cries from laboratory workers that their overnight bacterial cultures...