P1 phage

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Escherichia virus P1
Virus classification OOjs UI icon edit-ltr.svg
(unranked): Virus
Realm: Duplodnaviria
Kingdom: Heunggongvirae
Phylum: Uroviricota
Class: Caudoviricetes
Order: Caudovirales
Family: Myoviridae
Genus: Punavirus
Species:
Escherichia virus P1

P1 is a temperate bacteriophage that infects Escherichia coli and some other bacteria. When undergoing a lysogenic cycle the phage genome exists as a plasmid in the bacterium [1] unlike other phages (e.g. the lambda phage) that integrate into the host DNA. P1 has an icosahedral head containing the DNA attached to a contractile tail with six tail fibers. The P1 phage has gained research interest because it can be used to transfer DNA from one bacterial cell to another in a process known as transduction. As it replicates during its lytic cycle it captures fragments of the host chromosome. If the resulting viral particles are used to infect a different host the captured DNA fragments can be integrated into the new host's genome. This method of in vivo genetic engineering was widely used for many years and is still used today, though to a lesser extent. P1 can also be used to create the P1-derived artificial chromosome cloning vector which can carry relatively large fragments of DNA. P1 encodes a site-specific recombinase, Cre, that is widely used to carry out cell-specific or time-specific DNA recombination by flanking the target DNA with loxP sites (see Cre-Lox recombination).

Contents

Morphology

The virion is similar in structure to the T4 phage but simpler. [1] It has an icosahedral head [2] containing the genome attached at one vertex to the tail. The tail has a tube surrounded by a contractile sheath. It ends in a base plate with six tail fibres. The tail fibres are involved in attaching to the host and providing specificity. [3]

Genome

The genome of the P1 phage is moderately large, around 93Kbp [1] in length (compared to the genomes of e.g. T4 - 169Kbp, lambda - 48Kbp and Ff - 6.4Kbp). In the viral particle it is in the form of a linear double stranded DNA molecule. Once inserted into the host it circularizes and replicates as a plasmid. [4] [5]

In the viral particle the DNA molecule is longer (110Kbp) than the actual length of the genome. It is created by cutting an appropriately sized fragment from a concatemeric DNA chain having multiple copies of the genome (see the section below on lysis for how this is made). Due to this the ends of the DNA molecule are identical. This is referred to as being terminally redundant. This is important for the DNA to be circularized in the host. Another consequence of the DNA being cut out of a concatemer is that a given linear molecule can start at any location on the circular genome. This is called a cyclic permutation. [4]

The genome is especially rich in Chi sequences recognized by the bacterial recombinase RecBCD. The genome contains two origins of replication: oriR which replicates it during the lysogenic cycle and oriL which replicates it during the lytic stage. The genome of P1 encodes three tRNAs which are expressed in the lytic stage. [1]

Proteome. The genome of P1 encodes 112 proteins and 5 untranslated genes and is this about twice the size of bacteriophage lambda. [1]

Life cycle

Infection and early stages

The phage particle adsorbs onto the surface of the bacterium using the tail fibers for specificity. The tail sheath contracts and the DNA of the phage is injected into the host cell. The host DNA recombination machinery or the cre enzyme translated from the viral DNA recombine the terminally redundant ends and circularize the genome. Depending on various physiological cues, the phage may immediately proceed to the lytic phase or it may enter a lysogenic state. [5]

The gene that encodes the tail fibers have a set of sequences that can be targeted by a site specific recombinase Cin. This causes the C terminal end of the protein to switch between two alternate forms at a low frequency. The viral tail fibers are responsible for the specificity of binding to the host receptor. The targets of the viral tail fibers are under a constant pressure to evolve and evade binding. This method of recombinational diversity of the tail allows the virus to keep up with the bacterium. [6] This system has close sequence homologies to recombinational systems in the tail fibers of unrelated phages like the mu phage and the lambda phage.

Lysogeny

The genome of the P1 phage is maintained as a low copy number plasmid in the bacterium. The relatively large size of the plasmid requires [1] it to keep a low copy number lest it become too large a metabolic burden while it is a lysogen. As there is usually only one copy of the plasmid per bacterial genome, the plasmid stands a high chance of not being passed to both daughter cells. [5] The P1 plasmid combats this by several methods:

Lysis

The P1 plasmid has a separate origin of replication (oriL) that is activated during the lytic cycle. Replication begins by a regular bidirectional theta replication at oriL but later in the lytic phase, it switches to a rolling circle method of replication using the host recombination machinery. [1] [11] [12] This results in numerous copies of the genome being present on a single linear DNA molecule called a concatemer. The end of the concatemer is cut a specific site called the pac site or packaging site. [13] This is followed by the packing of the DNA into the heads till they are full. The rest of the concatemer that does not fit into one head is separated and the machinery begins packing this into a new head. The location of the cut is not sequence specific. Each head holds around 110kbp of DNA [13] so there is a little more than one complete copy of the genome (~90kbp) in each head, with the ends of the strand in each head being identical. After infecting a new cell this terminal redundancy is used by the host recombination machinery to cyclize the genome if it lacks two copies of the lox locus. [1] [13] If two lox sites are present (one in each terminally redundant end) the cyclization is carried out by the Cre recombinase. [1] [14]

Once the complete virions are assembled, the host cell is lysed, releasing the viral particles. [15]

History

P1 was discovered in 1951 by Giuseppe Bertani in Salvador Luria's laboratory, but the phage was little studied until Ed Lennox, also in Luria's group, showed in 1954–5 that it could transduce genetic material between host bacteria. This discovery led to the phage being used for genetic exchange and genome mapping in E. coli, and stimulated its further study as a model organism. [1] [16] [17] In the 1960s, Hideo Ikeda and Jun-ichi Tomizawa showed the phage's DNA genome to be linear and double-stranded, with redundancy at the ends. In the 1970s, Nat Sternberg characterised the Cre–lox site-specific recombination system, which allows the linear genome to circularise to form a plasmid after infection. During the 1980s, Sternberg developed P1 as a vector for cloning large pieces of eukaryotic DNA. [16] A P1 gene map based on a partial DNA sequence was published in 1993 by Michael Yarmolinsky and Małgorzata Łobocka, and the genome was completely sequenced by Łobocka and colleagues in 2004. [1] [17]

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<span class="mw-page-title-main">Lambda phage</span> Bacteriophage that infects Escherichia coli

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<span class="mw-page-title-main">Cloning vector</span> Small piece of maintainable DNA

A cloning vector is a small piece of DNA that can be stably maintained in an organism, and into which a foreign DNA fragment can be inserted for cloning purposes. The cloning vector may be DNA taken from a virus, the cell of a higher organism, or it may be the plasmid of a bacterium. The vector contains features that allow for the convenient insertion of a DNA fragment into the vector or its removal from the vector, for example through the presence of restriction sites. The vector and the foreign DNA may be treated with a restriction enzyme that cuts the DNA, and DNA fragments thus generated contain either blunt ends or overhangs known as sticky ends, and vector DNA and foreign DNA with compatible ends can then be joined by molecular ligation. After a DNA fragment has been cloned into a cloning vector, it may be further subcloned into another vector designed for more specific use.

<span class="mw-page-title-main">Prophage</span> Bacteriophage genome that is integrated into a bacterial cell

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<span class="mw-page-title-main">Lytic cycle</span> Cycle of viral reproduction

The lytic cycle is one of the two cycles of viral reproduction, the other being the lysogenic cycle. The lytic cycle results in the destruction of the infected cell and its membrane. Bacteriophages that only use the lytic cycle are called virulent phages.

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<span class="mw-page-title-main">Lysogenic cycle</span> Process of virus reproduction

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Cre recombinase is a tyrosine recombinase enzyme derived from the P1 bacteriophage. The enzyme uses a topoisomerase I-like mechanism to carry out site specific recombination events. The enzyme (38kDa) is a member of the integrase family of site specific recombinase and it is known to catalyse the site specific recombination event between two DNA recognition sites. This 34 base pair (bp) loxP recognition site consists of two 13 bp palindromic sequences which flank an 8bp spacer region. The products of Cre-mediated recombination at loxP sites are dependent upon the location and relative orientation of the loxP sites. Two separate DNA species both containing loxP sites can undergo fusion as the result of Cre mediated recombination. DNA sequences found between two loxP sites are said to be "floxed". In this case the products of Cre mediated recombination depends upon the orientation of the loxP sites. DNA found between two loxP sites oriented in the same direction will be excised as a circular loop of DNA whilst intervening DNA between two loxP sites that are opposingly orientated will be inverted. The enzyme requires no additional cofactors or accessory proteins for its function.

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References

  1. 1 2 3 4 5 6 7 8 9 10 11 12 Łobocka, Małgorzata B.; Debra J. Rose; Guy Plunkett; Marek Rusin; Arkadiusz Samojedny; Hansjörg Lehnherr; Michael B. Yarmolinsky; Frederick R. Blattner (November 2004). "Genome of Bacteriophage P1". Journal of Bacteriology. 186 (21): 7032–7068. doi:10.1128/JB.186.21.7032-7068.2004. ISSN   0021-9193. PMC   523184 . PMID   15489417.
  2. ↑ Walker, J T; D H Walker (March 1983). "Coliphage P1 morphogenesis: analysis of mutants by electron microscopy". Journal of Virology. 45 (3): 1118–1139. doi:10.1128/JVI.45.3.1118-1139.1983. ISSN   0022-538X. PMC   256520 . PMID   6834479.
  3. ↑ Huan, Yang W.; Fa-arun, Jidapha; Wang, Baojun (2022-11-15). "The Role of O-antigen in P1 Transduction of Shigella flexneri and Escherichia coli with its Alternative S' Tail Fibre". Journal of Molecular Biology. 434 (21): 167829. doi: 10.1016/j.jmb.2022.167829 . ISSN   0022-2836. PMID   36116540. S2CID   252325855.
  4. 1 2 Fujisawa, Hisao; Morita, Miyo (1997). "Phage DNA packaging". Genes to Cells. 2 (9): 537–545. doi:10.1046/j.1365-2443.1997.1450343.x. ISSN   1356-9597. PMID   9413995. S2CID   26780439.
  5. 1 2 3 Zhang, Kailun; Pankratz, Kiara; Duong, Hau; Theodore, Matthew; Guan, Jingwen; Jiang, Anxiao (Andrew); Lin, Yiruo; Zeng, Lanying (2021). "Interactions between Viral Regulatory Proteins Ensure an MOI-Independent Probability of Lysogeny during Infection by Bacteriophage P1". mBio. 12 (5): e01013–21. doi:10.1128/mBio.01013-21. ISSN   2150-7511. PMC   8546580 . PMID   34517752.
  6. ↑ Sandmeier, H.; S. Iida; W. Arber (1992-06-01). "DNA Inversion Regions Min of Plasmid p15B and Cin of Bacteriophage P1: Evolution of Bacteriophage Tail Fiber Genes". Journal of Bacteriology. 174 (12): 3936–3944. doi:10.1128/jb.174.12.3936-3944.1992. ISSN   0021-9193. PMC   206102 . PMID   1534556.
  7. ↑ Adams, David E.; James B. Bliska; Nicholas R. Cozzarelli (1992-08-05). "Cre-lox recombination in Escherichia coli cells mechanistic differences from the in Vitro reaction". Journal of Molecular Biology. 226 (3): 661–673. doi:10.1016/0022-2836(92)90623-R. ISSN   0022-2836. PMID   1324323.
  8. ↑ Austin, S; M Ziese; N Sternberg (September 1981). "A novel role for site-specific recombination in maintenance of bacterial replicons". Cell. 25 (3): 729–736. doi:10.1016/0092-8674(81)90180-X. ISSN   0092-8674. PMID   7026049. S2CID   28785596.
  9. ↑ Lehnherr, Hansjörg; Maguin, Emmanuelle; Jafri, Samina; Yarmolinsky, Michael B. (1993-10-05). "Plasmid Addiction Genes of Bacteriophage P1: doc, which Causes Cell Death on Curing of Prophage, and phd, which Prevents Host Death when Prophage is Retained". Journal of Molecular Biology. 233 (3): 414–428. doi:10.1006/jmbi.1993.1521. ISSN   0022-2836. PMID   8411153.
  10. ↑ Gazit, Ehud; Sauer, Robert T. (1999). "The Doc Toxin and Phd Antidote Proteins of the Bacteriophage P1 Plasmid Addiction System Form a Heterotrimeric Complex". Journal of Biological Chemistry. 274 (24): 16813–16818. doi: 10.1074/jbc.274.24.16813 . ISSN   0021-9258. PMID   10358024.
  11. ↑ Cohen, Gerald; Etti Or; Wolfgang Minas; Nat L. Sternberg (1996-10-10). "The bacteriophage P 1 lytic replicon: directionality of replication and cis-acting elements". Gene. 175 (1–2): 151–155. doi:10.1016/0378-1119(96)00141-2. ISSN   0378-1119. PMID   8917092.
  12. ↑ Cohen, Gerald (November 1983). "Electron microscopy study of early lytic replication forms of bacteriophage P1 DNA". Virology. 131 (1): 159–170. doi:10.1016/0042-6822(83)90542-1. ISSN   0042-6822. PMID   6359666.
  13. 1 2 3 Sternberg, N.; J. Coulby (1990-10-01). "Cleavage of the Bacteriophage P1 Packaging Site (pac) Is Regulated by Adenine Methylation". Proceedings of the National Academy of Sciences. 87 (20): 8070–8074. Bibcode:1990PNAS...87.8070S. doi: 10.1073/pnas.87.20.8070 . ISSN   0027-8424. PMC   54894 . PMID   2236019.
  14. ↑ Sternberg, N.; Sauer, B.; Hoess, R.; Abremski, K. (1986-01-20). "Bacteriophage P1 cre gene and its regulatory region: Evidence for multiple promoters and for regulation by DNA methylation". Journal of Molecular Biology. 187 (2): 197–212. doi:10.1016/0022-2836(86)90228-7. ISSN   0022-2836. PMID   3486297.
  15. ↑ Bednarek, Agnieszka; Cena, Agata; Izak, Wioleta; Bigos, Joanna; Łobocka, Małgorzata (2022). "Functional Dissection of P1 Bacteriophage Holin-like Proteins Reveals the Biological Sense of P1 Lytic System Complexity". International Journal of Molecular Sciences. 23 (8): 4231. doi: 10.3390/ijms23084231 . ISSN   1422-0067. PMC   9025707 . PMID   35457047.
  16. 1 2 Michael Yarmolinsky; Ronald Hoess (2015), "The Legacy of Nat Sternberg: The Genesis of Cre-lox Technology", Annual Review of Virology , 2 (1): 25–40, doi:10.1146/annurev-virology-100114-054930, PMID   26958905
  17. 1 2 Hansjörg Lehnherr (2006), "Bacteriophage P1", in Richard Calendar (ed.), The Bacteriophages, Oxford University Press, p. 350, ISBN   0195148509
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