title: Herpesvirus quiescence (QIF) in neuronal cells VI: Correlative analysis demonstrates usefulness of QIF-PC12 cells to examine HSV-1 latency, reactivation and genes implicated in its regulation.
abstract: OBJECTIVE
  To compare the usefulness of the in vitro quiescently infected (QIF)-PC12 cell model(30) with the in vivo rabbit eye model of latency for the study of herpes simplex virus (HSV) genes implicated in reactivation from latency.
  
  
  METHODS
  HSV-1 strains 17+/pR20.5/5 and 17+/pR20.5/5/LAT, that were previously constructed by insertion of genes encoding beta-galactosidase, green fluorescent protein (GFP) or the latency associated transcript (LAT) open reading frame in the U(S)5 region,(34) were used to examine viral growth and inducible reactivation in the two models.
  
  
  RESULTS
  17+/pR20.5/5 exhibited diminished reactivation phenotype when compared with wild type 17+ in neuronal cells (i.e., QIF-PC12 cell model) and the rabbit eye model of latency. 17+/pR20.5/5/LAT, which contains the deregulated LAT gene, reactivated at wild type levels. Analysis of growth in neurally differentiated (ND)-PC12 cells demonstrated a low proportion of QIF cells expressed virus-encoded signals during the quiescent infection and a direct relationship between lytic viral growth in neuronal cells and reactivation phenotype. Even though 17+/pR20.5/5/LAT produced a more severe acute infection in the rabbit cornea, the different reactivation efficiency of 17+/pR20.5/5 and 17+/pR20.5/5/LAT in vivo and in vitro was not attributed to different viral genome copy number in the cells harboring cryptic genomes.
  
  
  CONCLUSIONS
  We conclude that 1) viral growth in neuronal cells correlates with reactivation phenotype in vivo and in vitro, 2) 17+/pR20.5/5 is attenuated in viral growth and reactivation in both models, and 3) 17+/pR20.5/5/LAT demonstrates wild-type phenotype for reactivation in both models. Attenuation of 17+/pR20.5/5 could be the result of the disruption of U(S)5 or a second site mutation. If the attenuation is the result of U(S)5 disruption, a gene that provides anti-apoptotic functions,( 41,42) this attenuation is more than compensated for by the expression of the LAT ORF. Overall, the findings indicate that the QIF-PC12 cell model is useful for segregating phases of reactivation, and particularly studying the inductive events involved in reactivation of a cryptic viral genome in neurally differentiated cells.
authors.affiliation: Oral Medicine Section, Department of Oral Health Practice and Department of Microbiology, Immunology & Molecular Genetics, University of Kentucky College of Dentistry and College of Medicine, Kentucky, Lexington, USA.
authors.forename: Craig
authors.initials: C
authors.lastname: Miller
authors.forename: Partha
authors.initials: P
authors.lastname: Bhattacharjee
authors.forename: Shiro
authors.initials: S
authors.lastname: Higaki
authors.forename: Robert
authors.initials: R
authors.lastname: Jacob
authors.forename: Robert
authors.initials: R
authors.lastname: Danaher
authors.forename: Hilary
authors.initials: H
authors.lastname: Thompson
authors.forename: James
authors.initials: J
authors.lastname: Hill
mesh_terms: D000818:Animals; D002455:Cell Division; D002460:Cell Line; D004279:DNA, Viral / Q000378:metabolism; D015870:Gene Expression; D015967:Gene Expression Regulation, Viral; D049452:Green Fluorescent Proteins; D018259:Herpesvirus 1, Human / Q000235:genetics / Q000502:physiology*; D016849:Keratitis, Herpetic / Q000821:virology*; D008164:Luminescent Proteins / Q000235:genetics / Q000378:metabolism; D035683:MicroRNAs; D009474:Neurons / Q000378:metabolism / Q000473:pathology / Q000821:virology*; D016716:PC12 Cells / Q000821:virology; D011817:Rabbits; D051381:Rats; D014764:Viral Proteins / Q000235:genetics; D014775:Virus Activation / Q000235:genetics; D017735:Virus Latency / Q000235:genetics*; D001616:beta-Galactosidase / Q000235:genetics / Q000378:metabolism
chemical_list: D004279:DNA, Viral; D008164:Luminescent Proteins; D035683:MicroRNAs; D014764:Viral Proteins; C477253:latency associated transcript, herpes simplex virus-1; D049452:Green Fluorescent Proteins; D001616:beta-Galactosidase