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.
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 micrograms/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-synthesized 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 2 alpha (eIF-2 alpha) protein kinase DAI (p68), and that this property is dependent on the secondary structure of the small RNA molecule.
Clarke
P A
PA
Department of Cellular and Molecular Sciences, St. George's Hospital Medical School, London, England.
Sharp
N A
NA
Clemens
M J
MJ
D000818:Animals; D001483:Base Sequence; D002474:Cell-Free System; D004854:Herpesvirus 4, Human / Q000235:genetics*; D007700:Kinetics; D007930:Leucine / Q000378:metabolism; D008715:Methionine / Q000378:metabolism; D008969:Molecular Sequence Data; D015345:Oligonucleotide Probes; D010957:Plasmids; D011070:Poly I-C / Q000494:pharmacology*; D014176:Protein Biosynthesis / Q000187:drug effects; D012330:RNA, Double-Stranded / Q000494:pharmacology*; D012333:RNA, Messenger / Q000235:genetics*; D012367:RNA, Viral / Q000235:genetics*; D015183:Restriction Mapping; D012156:Reticulocytes / Q000378:metabolism*; D014158:Transcription, Genetic
D015345:Oligonucleotide Probes; D012330:RNA, Double-Stranded; D012333:RNA, Messenger; D012367:RNA, Viral; D008715:Methionine; D007930:Leucine; D011070:Poly I-C