Translational control by the Epstein‐Barr virus small RNA EBER‐1: Reversal of the double‐stranded RNA‐induced inhibition of protein synthesis in reticulocyte lysates

Abstract

A role for the Epstein‐Barr virus small RNA species EBER‐1 in the regulation of protein synthesis has been investigated in the reticulocyte‐lysate cell‐free translation system. Recombinant EBER‐1 was synthesized by in vitro transcription of a plasmid containing the viral gene and purified by CF11‐cellulose chromatography and ribonuclease III treatment. When added to the reticulocyte lysate at 10–20 μg/ml or more, EBER‐1 prevents the inhibition of protein synthesis caused by low concentrations of synthetic double‐stranded RNA, poly(I) · poly(C). This effect is eliminated by treatment of the recombinant EBER‐1 with ribonuclease T1. Disruption of the secondary structure of EBER‐1 by substitution of inosine for guanosine in the in‐vitro‐syntheized RNA impairs the ability of EBER‐1 to prevent the poly(I) · poly(C)‐mediated inhibition of protein synthesis. These results suggest that high concentrations of EBER‐1 regulate protein synthesis by blocking the activation of the double‐stranded RNA‐dependent eukaryotic initiation factor 2α (eIF‐2α) protein kinase DAI (p68), and that this property is dependent on the secondary structure of the small RNA molecule. Copyright © 1990, Wiley Blackwell. All rights reserved