Translational requirement of La Crosse virus S-mRNA synthesis: in vitro studies

Abstract

The exceptional requirement of La Crosse virus mRNA synthesis for ongoing protein synthesis in vivo was examined in vitro by using purified virions and a reticulocyte lysate. Transcription from the S genome produced two incomplete transcripts (110 and 205 nucleotides [nt]) in the absence of the lysate, whereas S-mRNA (900 nt) was predominantly made when the lysate was present. The addition of drugs which inhibit protein synthesis also inhibited the synthesis of S-mRNA, and in some cases led to the reappearance of the 205-nt RNA. Reconstruction experiments demonstrated that the incomplete transcripts were not the result of rapid and selective degradation of S-mRNA but were due to premature termination of the polymerase at defined sites. The requirement for ongoing protein synthesis for productive transcription in vitro is not at the level of chain initiation but for elongation of the nascent RNA beyond these sites.