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. 2001 Feb;75(4):1681-8.
doi: 10.1128/JVI.75.4.1681-1688.2001.

Fowlpox virus encodes a novel DNA repair enzyme, CPD-photolyase, that restores infectivity of UV light-damaged virus

V Srinivasan  1 , W M Schnitzlein, D N Tripathy
Affiliations

Fowlpox virus encodes a novel DNA repair enzyme, CPD-photolyase, that restores infectivity of UV light-damaged virus

V Srinivasan et al. J Virol. 2001 Feb.

Abstract

Fowlpox virus (FPV), a pathogen of poultry, can persist in desiccated scabs shed from infected hosts. Although the mechanisms which ensure virus survival are unknown, it is likely that some type of remedial action against environmentally induced damage is required. In this regard, we have identified an open reading frame (ORF) coding for a putative class II cyclobutane pyrimidine dimer (CPD)-photolyase in the genome of FPV. This enzyme repairs the UV light-induced formation of CPDs in DNA by using blue light as an energy source and thus could enhance the viability of FPV during its exposure to sunlight. Based on transcriptional analyses, the photolyase gene was found to be expressed late during the FPV replicative cycle. That the resultant protein retained DNA repair activity was demonstrated by the ability of the corresponding FPV ORF to complement functionally a photolyase-deficient Escherichia coli strain. Interestingly, insertional inactivation of the FPV photolyase gene did not impair the replication of such a genetically altered virus in cultured cells. However, greater sensitivity of this mutant than of the parental virus to UV light irradiation was evident when both were subsequently photoreactivated in the absence of host participation. Therefore, FPV appears to incorporate its photolyase into mature virions where the enzyme can promote their survival in the environment. Although expression of a homologous protein has been predicted for some chordopoxviruses, this report is the first to demonstrate that a poxvirus can utilize light to repair damage to its genome.

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Figures

FIG. 1
FIG. 1
Multiple amino aid sequence alignments of poxvirus and opossum photolyases. The deduced amino acid sequences of photolyases of FPV (accession no. AF246697) myxoma virus (MXV; accession no. AAF15015), Shope fibroma virus (SFV; accession no. AAF18004), M. sanguinipes entomopox virus (MSV; accession no. AAC97743), and short-tailed opossum (Monodelphis domestica) (STP; accession no. S50083) were aligned with the Clustal W program and analyzed for the presence of specific motifs in the Prosite database. Regions of conserved amino acids are shaded, and locations of the class II DNA photolyase signature sequences (PS01083 and PS01084) are underlined.
FIG. 2
FIG. 2
Phylogenetic tree of class II DNA photolyases. The amino acid sequences of vertebrate and invertebrate photolyases were aligned by using the Clustal W program. The alignment was manually edited, and a phylogenetic tree was generated by the maximum-likelihood method. Sources of enzymes: fruit fly (Drosophila melanogaster), accession no. S52047; goldfish (Carassius auratus), accession no. P34205; Japanese medaka (Oryzias latipes), accession no. BAA05043; long-nosed potoroo (Potorous tridactylus), accession no. BAA05041; short-tailed opossum (Monodelphis domestica), accession no. S50083; entomopox virus, accession no. AAC97743; fowlpox virus, accession no. AF246697; Shope fibroma virus, accession no. AAF18004; myxoma virus, accession no. AAF15015; Chlamydomonas reinharditii, accession no. AAD39433; Methanobacterium thermoautotrophicum, accession no. P12769; Myxococcus xanthus, accession no. AAC43723; Mycobacterium smegmatis, accession no. AAF04135; cyanobacterium (Anacystis nidulans), accession no. P05327; Bacillus firmis, accession no. 2017201A; thale cress (Arabidopsis thaliana), accession no. BAA74701.
FIG. 3
FIG. 3
Requirement of DNA replication for expression of the FPV photolyase gene. (A) RNAs extracted from FPV-infected QT-35 cells in the presence (lanes 1, 3, and 5) or absence of (lanes 2, 4, and 6) of cytosine arabinoside were subjected to RT-PCR using primers specific for the FPV thymidine kinase (lanes 1 and 2), A-type inclusion body protein (lanes 3 and 4), or photolyase (lanes 5 and 6) gene transcript. (B) RNAs extracted from FPV-infected QT-35 cells were reverse transcribed in the presence of random hexamer primers and then subjected to PCR using primers specific for either the FPV photolyase gene transcript (lane 2) or potential read-through transcripts encompassing a region including the photolyase gene promoter and 29% of the downstream coding sequence (lane 3). The extracted RNAs as well as virion DNA were also directly subjected to PCR using either the photolyase gene (lane 1) or read-through (lane 4) transcript-specific primer. The amplicons were electrophoresed in a 1.0% agarose gel, and their sizes (in base pairs) are indicated at the right.
FIG. 4
FIG. 4
Phenotypic complementation of photolyase-deficient E. coli. Plasmid pPHR1 was used to transform E. coli strain CSR 603. Exponentially growing cultures of the resultant bacterium (Phr) were kept for 3 h in the presence of arabinose (Ara) or glucose (Glu) to induce or repress, respectively, expression of the introduced FPV photolyase gene. Diluted cultures were then exposed to the indicated amounts of UV light and subsequently photoreactivated (PR) or kept in darkness (dark) for 1 h at ambient conditions. Survivability was based on CFU of the treated bacterial suspensions relative to CFU of the original stocks. For comparison, a UV inactivation curve of the parental, untransformed bacterium which had been placed in arabinose-containing medium and photoreactivated (CSR 603) is shown. The data represent the average of three independent determinations; standard deviations are indicated as error bars.
FIG. 5
FIG. 5
Growth kinetics of parental and photolyase-deficient recombinant FPV. QT-35 cell monolayers were infected at 0.1 PFU/cell with parental (Phr+) or recombinant (Phr−) FPV. At the indicated times, the cells and the medium were frozen and titered for the amount of infectious virus. The data represent the average of two independent determinations; standard deviations are indicated as error bars.
FIG. 6
FIG. 6
UV light inactivation of parental and photolyase-deficient recombinant FPV. Extracellular parental (Phr+) and recombinant (Phr−) virions were exposed to the indicated doses of UV light and then either photoreactivated (PR) or kept in darkness (dark) for 1 h at ambient conditions. Survivability was based on titers of the treated virus suspensions relative to titers of the original stocks. The results represent the average of three independent determinations; standard deviations are indicated as error bars.
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References

    1. Adachi J, Hasegawa M. Molphy version 2.3 programs for molecular phylogenetics based on maximum likelihood. Computer Science Monographs no. 28. Tokyo, Japan: Institute of Statistical Methods; 1996.
    1. Afonso C L, Tulman E R, Lu Z, Oma E, Kutish G F, Rock D L. The genome of Melanoplus sanguinipesentomopox virus. J Virol. 1999;73:533–552. - PMC - PubMed
    1. Afonso C L, Tulman E R, Lu Z, Zsak L, Kutish G F, Rock D L. The genome of fowlpox virus. J Virol. 2000;74:3815–3831. - PMC - PubMed
    1. Altschul S F, Madden T L, Schaeffer A A, Zhang J, Zhang Z, Miller W, Lipman D J. Gapped BLAST and PSI-BLAST: a new generation of protein database search programs. Nucleic Acids Res. 1997;25:3389–3402. - PMC - PubMed
    1. Baldick Jr C J, Keck J G, Moss B. Mutational analysis of the core, spacer, and initiator regions of vaccinia virus intermediate-class promoters. J Virol. 1992;66:4710–4719. - PMC - PubMed

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