Reversal of the double-stranded-RNA-induced inhibition of protein synthesis by a catalytically inactive mutant of the protein kinase PKR.
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.
Sharp
T V
TV
Department of Cellular and Molecular Sciences, St George's Hospital Medical School, London, England.
Xiao
Q
Q
Jeffrey
I
I
Gewert
D R
DR
Clemens
M J
MJ
D000818:Animals; D016367:Baculoviridae / Q000235:genetics; D002478:Cells, Cultured; D009036:Moths / Q000166:cytology; D017354:Point Mutation; D014176:Protein Biosynthesis; D017346:Protein Serine-Threonine Kinases / Q000235:genetics* / Q000378:metabolism*; D012330:RNA, Double-Stranded / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism; D019892:eIF-2 Kinase
D012330:RNA, Double-Stranded; D011994:Recombinant Proteins; D017346:Protein Serine-Threonine Kinases; D019892:eIF-2 Kinase