Vaccinia virus (VV), the prototypic poxvirus, encodes a repertoire of
proteins responsible for the metabolism of its large dsDNA genome. Previous work has furthered our understanding of how poxviruses replicate and recombine their genomes, but little is known about whether the poxvirus genome undergoes DNA repair. Our studies here are aimed at understanding how VV responds to exogenous DNA damage introduced by UV irradiation. Irradiation of cells prior to
infection decreased
protein synthesis and led to an ∼12-fold reduction in viral yield. On top of these cell-specific insults, irradiation of VV
infections at 4 h postinfection (hpi) introduced both cyclobutene
pyrimidine dimer (CPD) and 6,4-photoproduct (6,4-PP) lesions into the viral genome led to a nearly complete halt to further
DNA synthesis and to a further reduction in viral yield (∼35-fold). DNA lesions persisted throughout
infection and were indeed present in the genomes encapsidated into nascent virions. Depletion of several cellular
proteins that mediate nucleotide excision repair (XP-A, -F, and -G) did not render
viral infections hypersensitive to UV. We next investigated whether
viral proteins were involved in combatting DNA damage.
Infections performed with a virus lacking the A50
DNA ligase were moderately hypersensitive to UV irradiation (∼3-fold). More strikingly, when the
DNA polymerase inhibitor cytosine arabinoside (araC) was added to wild-type
infections at the time of UV irradiation (4 hpi), an even greater
hypersensitivity to UV irradiation was seen (∼11-fold). Virions produced under the latter condition contained elevated levels of CPD adducts, strongly suggesting that the viral polymerase contributes to the repair of UV lesions introduced into the viral genome. IMPORTANCE Poxviruses remain of significant interest because of their continuing clinical relevance, their utility for the development of
vaccines and oncolytic
therapies, and their illustration of fundamental principles of viral replication and virus/cell interactions. These viruses are unique in that they replicate exclusively in the cytoplasm of infected mammalian cells, providing novel challenges for DNA viruses. How poxviruses replicate, recombine, and possibly repair their genomes is still only partially understood. Using UV irradiation as a form of exogenous DNA damage, we have examined how vaccinia virus metabolizes its genome following insult. We show that even UV irradiation of cells prior to
infection diminishes viral yield, while UV irradiation during
infection damages the genome, causes a halt in
DNA accumulation, and reduces the viral yield more severely. Furthermore, we show that
viral proteins, but not the cellular machinery, contribute to a partial repair of the viral genome following UV irradiation.