Reversal of the double‐stranded‐RNA‐induced inhibition of protein synthesis by a catalytically inactive mutant of the protein kinase PKR

Abstract

The interferon‐inducible double‐stranded‐RNA(dsRNA)‐dependent protein kinase PKR has been implicated in both the antiviral and cell growth‐regulatory effects of the interferons. Over‐expression of the wild‐type form of this protein inhibits cell proliferation, whereas over‐expression of inactive mutant forms transforms cells to a tumourigenic phenotype. It has been suggested that mutant PKR exerts a dominant negative effect on the activity of the wild‐type protein kinase. We have investigated this possibility using the rabbit reticulocyte cell‐free translation system in which protein synthesis is inhibited by dsRNA due to activation of PKR and phosphorylation of initiation factor eIF‐2. Addition of a highly purified inactive PKR mutant, synthesised in a baculovirus‐infected insect cell system, rescues protein synthesis from inhibition by low concentrations of dsRNA in a dose‐dependent manner. The PKR mutant has no effect on protein synthesis in the absence of dsRNA or in the presence of another inhibitory protein kinase, the haem‐controlled repressor. Inhibition of translation can be re‐established in the presence of the mutant PKR by adding a higher concentration of dsRNA. These results suggest that inactive mutant PKR does exert a dominant negative effect on wild‐type PKR and that this may be due to competition for dsRNA binding. Copyright © 1993, Wiley Blackwell. All rights reserved