Genomic Analysis of Pseudomonas putida Phage tf with Localized Single-Strand DNA Interruptions

Abstract

The complete sequence of the 46,267 bp genome of the lytic bacteriophage tf specific to Pseudomonas putida PpG1 has been determined. The phage genome has two sets of convergently transcribed genes and 186 bp long direct terminal repeats. The overall genomic architecture of the tf phage is similar to that of the previously described Pseudomonas aeruginosa phages PaP3, LUZ24 and phiMR299-2, and 39 out of the 72 products of predicted tf open reading frames have orthologs in these phages. Accordingly, tf was classified as belonging to the LUZ24-like bacteriophage group. However, taking into account very low homology levels between tf DNA and that of the other phages, tf should be considered as an evolutionary divergent member of the group. Two distinguishing features not reported for other members of the group were found in the tf genome. Firstly, a unique end structure – a blunt right end and a 4-nucleotide 3′-protruding left end – was observed. Secondly, 14 single-chain interruptions (nicks) were found in the top strand of the tf DNA. All nicks were mapped within a consensus sequence 5′-TACT/RTGMC-3′. Two nicks were analyzed in detail and were shown to be present in more than 90% of the phage population. Although localized nicks were previously found only in the DNA of T5-like and phiKMV-like phages, it seems increasingly likely that this enigmatic structural feature is common to various other bacteriophages.

The Susceptibility of Pseudomonas aeruginosa Strains from Cystic Fibrosis Patients to Bacteriophages

“…We observed a very important relative amount of sequencing reads at 8 positions along the genome and detected the consensus sequence GTACTATGAC at each of these positions. This sequence is very close to the consensus sequence 5′-TACTRTGMC-3′ corresponding to single-strand DNA interruptions in phage tf of P. putida [46]. Such nicks were never described in P. aeruginosa LUZ24-like phages although the site is present in all of the sequenced genomes reported so far.…”

Genomic hypervariability of phage Andromeda is unique among known dsDNA viruses

“…aeruginosa [2] and P. putida phage tf [31]. A search of the Andromeda genome failed to find the consensus associated with nicks in P. aeruginosa phages or P. putida phage.…”

The Essential Co-Option of Uracil-DNA Glycosylases by Herpesviruses Invites Novel Antiviral Design

“…Some retroviruses, such as HIV-1, apparently strategically manage uracil-DNA to efficiently integrate into host genomic DNA and must, therefore, silence as well as co-opt Ung [31,32]. Other viruses, as observed in unrelated lineages of bacteriophages, generate single-stranded DNA intermediates during replication or assembly [60][61][62][63]. Naked single-stranded DNA is at much higher risk of cytosine deamination than either protein-coated single strands [64], or duplex DNA [11].…”

“…In other types of virus, appreciable accumulation of DNA uracil in the wake of APOBEC activity will result in base-excision by Ung in close proximity on both strands of the affected DNA and thus pathogen DNA fragmentation due to BER-induced double-strand breaks (Figure 3b; Figure 4). In bacteria, Ung is present residually and is therefore a potent restriction enzyme against viral DNA that accumulates uracil as it is rapidly expanding (i.e., due to relatively lower fidelity of viral DNA polymerases and the effects of nucleotide pool bias) especially if utilising single-stranded intermediates [59][60][61][62][63]; as mentioned earlier, viruses employing uracil as a substitute for thymine are a prime target for Ung [38]. Ung acts as part of both the innate and humoral immune response pathways downstream of AID and APOBEC (Apolipoprotein B mRNA Editing Catalytic polypeptide-like family) enzymes, which enzymatically deaminate cytosine in DNA [65,66].…”