title: Absence of the uracil DNA glycosylase of murine gammaherpesvirus 68 impairs replication and delays the establishment of latency in vivo.
abstract: Uracil DNA glycosylases (UNG) are highly conserved proteins that preserve DNA fidelity by catalyzing the removal of mutagenic uracils. All herpesviruses encode a viral UNG (vUNG), and yet the role of the vUNG in a pathogenic course of gammaherpesvirus infection is not known. First, we demonstrated that the vUNG of murine gammaherpesvirus 68 (MHV68) retains the enzymatic function of host UNG in an in vitro class switch recombination assay. Next, we generated a recombinant MHV68 with a stop codon in ORF46/UNG (ΔUNG) that led to loss of UNG activity in infected cells and a replication defect in primary fibroblasts. Acute replication of MHV68ΔUNG in the lungs of infected mice was reduced 100-fold and was accompanied by a substantial delay in the establishment of splenic latency. Latency was largely, yet not fully, restored by an increase in virus inoculum or by altering the route of infection. MHV68 reactivation from latent splenocytes was not altered in the absence of the vUNG. A survey of host UNG activity in cells and tissues targeted by MHV68 indicated that the lung tissue has a lower level of enzymatic UNG activity than the spleen. Taken together, these results indicate that the vUNG plays a critical role in the replication of MHV68 in tissues with limited host UNG activity and this vUNG-dependent expansion, in turn, influences the kinetics of latency establishment in distal reservoirs.
  
  
  OBJECTIVE
  Herpesviruses establish chronic lifelong infections using a strategy of replicative expansion, dissemination to latent reservoirs, and subsequent reactivation for transmission and spread. We examined the role of the viral uracil DNA glycosylase, a protein conserved among all herpesviruses, in replication and latency of murine gammaherpesvirus 68. We report that the viral UNG of this murine pathogen retains catalytic activity and influences replication in culture. The viral UNG was impaired for productive replication in the lung. This defect in expansion at the initial site of acute replication was associated with a substantial delay of latency establishment in the spleen. The levels of host UNG were substantially lower in the lung compared to the spleen, suggesting that herpesviruses encode a viral UNG to compensate for reduced host enzyme levels in some cell types and tissues. These data suggest that intervention at the site of initial replicative expansion can delay the establishment of latency, a hallmark of chronic herpesvirus infection.
authors.affiliation: Department of Molecular Genetics and Microbiology, Stony Brook University, Stony Brook, New York, USA.
authors.forename: Nana
authors.initials: N
authors.lastname: Minkah
authors.affiliation: Department of Molecular Carcinogenesis, Science Park, The University of Texas MD Anderson Cancer Center, Smithville, Texas, USA.
authors.forename: Marc
authors.initials: M
authors.lastname: Macaluso
authors.affiliation: Gundersen Health System, La Crosse, Wisconsin, USA.
authors.forename: Darby G
authors.initials: DG
authors.lastname: Oldenburg
authors.affiliation: Department of Molecular Genetics and Microbiology, Stony Brook University, Stony Brook, New York, USA.
authors.forename: Clinton R
authors.initials: CR
authors.lastname: Paden
authors.affiliation: Gundersen Health System, La Crosse, Wisconsin, USA.
authors.forename: Douglas W
authors.initials: DW
authors.lastname: White
authors.affiliation: Department of Molecular Carcinogenesis, Science Park, The University of Texas MD Anderson Cancer Center, Smithville, Texas, USA.
authors.forename: Kevin M
authors.initials: KM
authors.lastname: McBride
authors.affiliation: Department of Molecular Genetics and Microbiology, Stony Brook University, Stony Brook, New York, USA laurie.krug@stonybrook.edu.
authors.forename: Laurie T
authors.initials: LT
authors.lastname: Krug
mesh_terms: D000818:Animals; D005260:Female; D015967:Gene Expression Regulation, Viral; D006566:Herpesviridae Infections / Q000662:veterinary* / Q000821:virology*; D051379:Mice; D008810:Mice, Inbred C57BL; D018144:Rhadinovirus / Q000201:enzymology* / Q000235:genetics / Q000502:physiology; D012376:Rodent Diseases / Q000821:virology*; D051981:Uracil-DNA Glycosidase / Q000172:deficiency* / Q000235:genetics; D017735:Virus Latency; D014779:Virus Replication
chemical_list: D051981:Uracil-DNA Glycosidase