Alphavirus minus-strand synthesis and persistence in mouse embryo fibroblasts derived from mice lacking RNase L and protein kinase R.
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.
Sawicki
Dorothea L
DL
Department of Microbiology and Immunology, Medical College of Ohio, Toledo, Ohio 43614, USA. dsawicki@mco.edu
Silverman
Robert H
RH
Williams
Bryan R
BR
Sawicki
Stanley G
SG
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*
D012367:RNA, Viral; D019892:eIF-2 Kinase; D004722:Endoribonucleases; C031956:2-5A-dependent ribonuclease