The antiviral enzymes PKR and RNase L suppress gene expression from viral and non-viral based vectors

Expression of transfected genes is shown to be suppressed by two intracellu lar enzymes, RNase L and protein kinase PKR, which function in interferon-t reated cells to restrict viral replication, RNase L-/- or PKR-/- murine emb ryonic fibroblasts produced enhanced levels of protein from transfected gen es compared with wild-type cells, Increased expression of exogenous genes i n RNase L-/- cells correlated with elevated levels of mRNA and thus appeare d to be due to enhanced mRNA stability. Plasmid encoding adenovirus VA RNAs was able to further enhance accumulation of the exogenous gene transcript and protein, even in cells lacking PKR, In contrast to the increased expres sion of transfected genes in cells lacking RNase L or PKR, expression of en dogenous host genes was unaffected by the absence of these enzymes. In addi tion, a dominant-negative PKR mutant improved expression from a conventiona l plasmid vector and from a Semliki Forest virus derived, self-replicating vector. These results indicate that viral infections and transfections prod uce similar stress responses in mammalian cells and suggest strategies for selectively increasing expression of exogenous genes.