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Phylogenetic structure and evolution of regulatory genes and integrases of P2-like phages
Stockholm University, Faculty of Science, Department of Genetics, Microbiology and Toxicology. (Team Nilsson)
Stockholm University, Faculty of Science, Department of Genetics, Microbiology and Toxicology. (Team Nilsson)
Stockholm University, Faculty of Science, Department of Genetics, Microbiology and Toxicology. (Team Nilsson)
Stockholm University, Faculty of Science, Department of Genetics, Microbiology and Toxicology. (Team Nilsson)
2011 (English)In: Bacteriophage, ISSN 2159-7073, Vol. 1, no 4, 207-218 p.Article in journal (Refereed) Published
Abstract [en]

The phylogenetic relationships and structural similarities of the proteins encoded within the regulatory region (containing the integrase gene and the lytic – lysogenic transcriptional switch genes) of P2-like phages were analyzed, and compared to the phylogenetic relationship of P2-like phages inferred from four structural genes. P2-like phages are thought to be one of the most genetically homogenous phage groups but the regulatory region nevertheless varies extensively between different phage genomes.

The analyses showed that there are many types of regulatory regions, but two types can be clearly distinguished; regions similar either to the phage P2 or to the phage 186 regulatory regions. These regions were also found to be most frequent among the sequenced P2-like phage or prophage genomes, and common in phages using Escherichia coli as a host. Both the phylogenetic and the structural analyses showed that these two regions are related. The integrases as well as the cox/apl genes show a common monophyletic origin but the immunity repressor genes, the type P2 C gene and the type 186 cI gene, are likely of different origin. There was no indication of recombination between the P2 – 186 types of regulatory genes but the comparison of the phylogenies of the regulatory region with the phylogeny based on four structural genes revealed recombinational events between the regulatory region and the structural genes.

Less common regulatory regions were phylogenetically heterogeneous and typically contained a fusion of genes from distantly related or unknown phages and P2-like genes.

Place, publisher, year, edition, pages
Austin, Texas, USA: Landes Bioscience , 2011. Vol. 1, no 4, 207-218 p.
Keyword [en]
Peduovirinae, P2-like bacteriophages, Phylogenetic analysis, Phage integration, Lytic-lysogenic transcriptional switch, Gamma-proteobacteria
National Category
Genetics Microbiology
Research subject
Molecular Genetics
Identifiers
URN: urn:nbn:se:su:diva-65365DOI: 10.4161/bact.1.4.18470OAI: oai:DiVA.org:su-65365DiVA: diva2:462929
Projects
The evolution of P2-like bacteriophages
Note
Article has also supplementary material, which is missing in this recordAvailable from: 2011-12-08 Created: 2011-12-08 Last updated: 2012-02-28Bibliographically approved
In thesis
1. Site-specific recombination in P2-like coliphages
Open this publication in new window or tab >>Site-specific recombination in P2-like coliphages
2009 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

The scope of these studies has been to investigate the site-specific recombination systems of P2-like coliphages, both in an evolutionary perspective by a comparative analysis of related phages as well as in a functional perspective.

Surveys of P2-like phages in Escherichia coli isolated from nature reveal the existence of seven discrete immunity classes and three integration sites, one of them previously unknown. Phylogenetic analysis of the repressor proteins and other analyses show that homologous recombination plays a role in the appearance of new immunities. Other studies of P2-like prophages from sequenced genomes from public databases show that the P2-like phages grow in different γ-proteobacteria. Based on the type of immunity and site-specific recombination system they can be roughly subdivided in two distinct subgroups and some new host integration sites could be identified. Some of the host attachment sites have a high identity to the sequences in the human genome, making them interesting as potential tools for targeted gene insertions into unmodified human cells.

The functional studies have been focused on the identification of the determinants for site specificity, which is important for the use of the enzyme for targeted gene insertions into unmodified genomes. Two approaches have been used. In one, we have performed a structure-function analysis of P2 Int that has identified several presumptive residues involved in specific binding to the core sequence, all of them located in the same alpha-helix. This knowledge could be a base for an in vitro evolution of the integrase to enable it to accept new DNA targets with a high affinity. With respect to the excisionases from P2-like coliphages integrating in different sites, we found that they share some common features when they bind and bend to their DNA targets, but there are also significant differences, especially those related to the number of binding sites and the distribution of these and the IHF binding sites in the attP regions. In the other approach we have started to characterize the site-specific recombination system of another P2-like phage, ΦD145, that has a host target with a high identity to a site in the human genome. This looks promising since the human sequence can be used in vivo in E. coli with a rather high efficiency.

Place, publisher, year, edition, pages
Stockholm: Institutionen för genetik, mikrobiologi och toxikologi, 2009. 41 p.
National Category
Genetics
Research subject
Molecular Genetics
Identifiers
urn:nbn:se:su:diva-8502 (URN)978-91-7155-817-6 (ISBN)
Public defence
2009-03-06, sal E306, Arrheniuslaboratorierna, Svante Arrhenius väg 14-18, Stockholm, 10:00 (English)
Opponent
Supervisors
Available from: 2009-02-12 Created: 2009-02-03 Last updated: 2012-02-28Bibliographically approved
2. A Genetic Switch in Bacteriophages within the Peduovirinae Subfamily: Structure, Function and Evolution
Open this publication in new window or tab >>A Genetic Switch in Bacteriophages within the Peduovirinae Subfamily: Structure, Function and Evolution
2012 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

The temperate bacteriophages in the Peduovirinae subfamily can either grow lytically or integrate into their bacterial host and form lysogeny. Which one of the two life cycles the phage will enter after infection is controlled by a transcriptional switch. The switch also controls the induction of genes necessary for an integrated phage, a prophage, to excise out of the host genome and propagate lytically. In its most simple form, the transcriptional switch consists of two proteins repressing each other’s promoters, which are oriented face to face in close proximity.

The Peduovirinae phages contain two types of transcriptional switches. They were studied with phylogenetic methods to determine their evolution and distribution. Bioinformatic analyses showed that there were several new E. coli integration sites and new inferred immunity classes among the Peduovirinae phages. The two switch types fell into two distinct groups, with no overlap in any of the proteins, but these groups were not defined by host barriers. But in vivo distribution did show a host preference.

The P2 C protein was crystallized and its 3D structure determined. It forms a symmetrical dimer in vitro, with an unstructured C-terminal end. The DNA binding domain was determined to lie in alpha helix three and narrowed down to three residues. The C terminal end of the protein is suggested to be part of tetramerization, but a nine amino acid truncation does not affect activity in vitro.

In an attempt to discover the mechanism between the switch from lysogeny to lysis in phage P2 the interactions between the two switch proteins and the proteins of its host E. coli was analyzed. Eight E. coli proteins interacted with protein C or Cox, but no interaction between the two switch proteins was detected. Two E. coli proteins showed a distinct effect on the expression of C, and several affected the level of phage lysis. The mechanisms behind these effects are still unclear.

Place, publisher, year, edition, pages
Stockholm: Department of Genetics, Microbiology and Toxicology, Stockholm University, 2012. 88 p.
National Category
Genetics
Research subject
Molecular Genetics
Identifiers
urn:nbn:se:su:diva-74031 (URN)978-91-7447-467-1 (ISBN)
Public defence
2012-03-30, Magnelisalen, Kemiska övningslaboratoriet, Svante Arrhenius väg 16 B, Stockholm, 13:00 (English)
Opponent
Supervisors
Note
At the time of the doctoral defence the following paper was unpublished and had a status as follows: Paper nr 3: ManuscriptAvailable from: 2012-03-08 Created: 2012-02-27 Last updated: 2012-03-01Bibliographically approved

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