The Genome and Structural Proteome of YuA, a New Pseudomonas aeruginosa Phage Resembling M6

Abstract

Pseudomonas aeruginosa phage YuA (Siphoviridae) was isolated from a pond near Moscow, Russia. It has an elongated head, encapsulating a circularly permuted genome of 58,663 bp, and a flexible, noncontractile tail, which is terminally and subterminally decorated with short fibers. The YuA genome is neither Mu-nor -like and encodes 78 gene products that cluster in three major regions involved in (i) DNA metabolism and replication, (ii) host interaction, and (iii) phage particle formation and host lysis. At the protein level, YuA displays significant homology with phages M6, JL001, 73, B3, DMS3, and D3112. Eighteen YuA proteins were identified as part of the phage particle by mass spectrometry analysis. Five different bacterial promoters were experimentally identified using a promoter trap assay, three of which have a 54 -specific binding site and regulate transcription in the genome region involved in phage particle formation and host lysis. The dependency of these promoters on the host 54 factor was confirmed by analysis of an rpoN mutant strain of P. aeruginosa PAO1. At the DNA level, YuA is 91% identical to the recently (July 2007) annotated phage M6 of the Lindberg typing set. Despite this level of DNA homology throughout the genome, both phages combined have 15 unique genes that do not occur in the other phage. The genome organization of both phages differs substantially from those of the other known Pseudomonas-infecting Siphoviridae, delineating them as a distinct genus within this family.Despite a decade of major sequencing efforts, many aspects of the genomic diversity among bacteriophages remain to be addressed. Recent metagenomic sequencing of uncultured viral communities from oceanic regions has shown that, although common patterns of genomic organization are present, up to 90% of marine-phage sequences are novel (6, 14). Other phage-sequencing projects revealed distinct levels of genomic diversity among phages infecting different bacteria. For example, the diversity of phage types infecting mycobacteria (37) contrasts sharply with dairy phages, which constitute a closeknit group (15).In recent years, genome sequencing efforts for phages infecting Pseudomonas aeruginosa have revealed this group as strongly diverse at the genome organizational level, which is consistent with their reported diversity in propagation, host interaction, and particle structure. The phages of P. aeruginosa are under investigation to determine the scope of their therapeutic potential and to unravel their dynamic interaction with their pathogenic host. Moreover, insight into the genome content of P. aeruginosa phages allows insight into the evolutionary aspects of these phages. At present, 27 complete genome sequences of phages infecting P. aeruginosa have been deposited in public databases (2). Among the siphoviruses infecting P. aeruginosa, phage D3112 is probably the best studied. With the exception of a DNA modification module and a structural region coding for tail morphogenesis proteins, phage D3112 shares its overall gen...

Deletion of σ 54 ( rpoN ) Alters the Rate of Autolysis and Biofilm Formation in Enterococcus faecalis

“…The role of 54 in regulating numerous biological properties, including those related to virulence, has been well documented for a variety of bacterial species (3,10,21,42,49,51,52,55,58,59). However, its role in E. faecalis has been limited to observations made regarding its contribution to sensitivity to class IIa bacteriocins through the regulation of sugar PTSs (9,13,22).…”

“…In P. aeruginosa, deletion of rpoN abolishes cell death in the microcolonies during biofilm maturation and has been related to the expression of surface structures (type 4 pili and flagella) whose expression is regulated by 54 (56). Additionally, 54 dependent gene regulation promotes phage-induced lysis in P. aeruginosa (10). There are seven phages associated with E. faecalis V583, with one of them being a part of the core genome (37).…”

Novel Phage Group Infecting Lactobacillus delbrueckii subsp. lactis, as Revealed by Genomic and Proteomic Analysis of Bacteriophage Ldl1

“…The structural protein profile was generated by standard Tris-glycine sodium dodecyl sulfate (SDS)-12% polyacrylamide gel electrophoresis (PAGE). Gel slices were then excised, trypsinized, and analyzed by using electrospray ionization tandem mass spectrometry (ESI-MS/MS), as previously described (28).…”

Molecular Characterization of Three Lactobacillus delbrueckii subsp. bulgaricus Phages

“…The structural protein profile was generated by standard Tris-glycine, sodium dodecyl sulfate (SDS)-12% polyacrylamide gel electrophoresis (PAGE) and visualized following staining with 0.25% Coomassie brilliant blue. Gel slices were then excised, trypsin treated, and analyzed using electrospray ionization-tandem mass spectrometry (ESI-MS/MS) as previously described (31,32).…”