title: Alphavirus minus-strand synthesis and persistence in mouse embryo fibroblasts derived from mice lacking RNase L and protein kinase R.
abstract: We report our studies to probe the possible role of the host response to double-stranded RNA in cessation of alphavirus minus-strand synthesis. Mouse embryo fibroblasts (MEF) from Mx1-deficient mice that also lack either the protein kinase R (PKR) or the latent RNase L or both PKR and RNase L were screened. In RNase L-deficient but not wild-type or PKR-deficient MEF, there was continuous synthesis of minus-strand templates and the formation of new replication complexes producing viral plus strands. Inhibiting translation caused minus-strand synthesis to stop and a loss of transcription activity of the mature replication complexes. This turnover of replication complexes that were stable in cells containing RNase L suggested that RNase L plays some role, albeit possibly indirect, in the formation of stable replication complexes during alphavirus infection. In addition, confluent monolayers of RNase L-deficient murine cells readily established persistent infections and were not killed. This phenotype is contrary to what has been observed for infection in vertebrate cells with a presumably functional RNase L gene and more resembled alphavirus replication in Aedes mosquito cells, in which the activity of replication complexes making plus stands was also found to decay with inhibition of translation.
authors.affiliation: Department of Microbiology and Immunology, Medical College of Ohio, Toledo, Ohio 43614, USA. dsawicki@mco.edu
authors.forename: Dorothea L
authors.initials: DL
authors.lastname: Sawicki
authors.forename: Robert H
authors.initials: RH
authors.lastname: Silverman
authors.forename: Bryan R
authors.initials: BR
authors.lastname: Williams
authors.forename: Stanley G
authors.initials: SG
authors.lastname: Sawicki
mesh_terms: D000818:Animals; D002642:Chick Embryo; D004722:Endoribonucleases / Q000235:genetics / Q000502:physiology*; D005347:Fibroblasts / Q000821:virology; D051379:Mice; D012367:RNA, Viral / Q000096:biosynthesis*; D012845:Sindbis Virus / Q000502:physiology*; D014779:Virus Replication; D019892:eIF-2 Kinase / Q000235:genetics / Q000502:physiology*
chemical_list: D012367:RNA, Viral; D019892:eIF-2 Kinase; D004722:Endoribonucleases; C031956:2-5A-dependent ribonuclease