Inhibition of protein synthesis and activation of nuclease in rabbit reticulocyte lysates by the unusual oligonucleotide pppA2'p5'A2'p5'A.

Abstract

The oligonucleotide 5′‐triphosphoadenylyl(2′‐5′)adenylyl(2′‐5′)adenosine (pppA2′p5′A2′p5′A) is synthesised by reticulocyte lysates and extracts from interferon‐treated cells in the presence of double‐stranded RNA. It inhibits the translation of both viral and non‐viral mRNAs and of poly‐(rU,rC) in the micrococcal‐nuclease‐treated reticulocyte lysate system. The translation of poly(rU) is insensitive in this assay, however. Inhibition develops in vitro after a lag period, the length of which depends on the nature of the mRNA present and its concentration.The degradation of radioactive Mengo virus RNA or L cell cytoplasmic RNA is accelerated in reticulocyte lysates incubated with the oligonucleotide. This is due to activation of one or more endonucleases which degrade(s) RNA to large oligonucleotide fragments. Our results suggest that the endonuclease is responsible for most (if not all) of the inhibitory effects of the oligonucleotide on protein synthesis in the reticulocyte lysate.In the presence of pppA2′p5′A2′p5′A, protein synthesis can be reactivated after it has ceased by addition of fresh mRNA. In contrast, initiation factor eIF‐2 is unable to prevent inhibition by the oligonucleotide under conditions where it protects the system from the effects of the haem‐controlled repressor and partially protects against inhibition by double‐stranded RNA.These findings are discussed in relation to the regulation of protein synthesis by double‐stranded RNA and by interferon.