Regulation of RNA synthesis by the genomic termini of vesicular stomatitis virus: identification of distinct sequences essential for transcription but not replication.
The RNA-dependent RNA polymerase of vesicular stomatitis virus (VSV), a nonsegmented negative-strand RNA virus, directs two discrete RNA synthetic processes, transcription and replication. Available evidence suggests that the two short extragenic regions at the genomic termini, the 3' leader (Le) and the complement of the 5' trailer (TrC), contain essential signals for these processes. We examined the roles in transcription and replication of sequences in Le and TrC by monitoring the effects of alterations to the termini of subgenomic replicons, or infectious viruses, on these RNA synthetic processes. Distinct elements in Le were found to be required for transcription that were not required for replication. The promoter for mRNA transcription was shown to include specific sequence elements within Le at positions 19 to 29 and 34 to 46, a separate element at nucleotides 47 to 50, the nontranscribed leader-N gene junction. The sequence requirements for transcription within the Le region could not be supplied by sequences found at the equivalent positions in TrC. In contrast, sequences from either Le or TrC functioned well to signal replication, indicating that within the confines of the VSV termini, the sequence requirements for replication were less stringent. Deletions engineered at the termini showed that the terminal 15 nucleotides of either Le or TrC allowed a minimal level of replication. Within these confines, levels of replication were affected by both the extent of complementarity between the genomic termini and the involvement of the template in transcription. In agreement with our previous observations, increasing the extent of complementarity between the natural termini increased levels of replication, and this effect was most operative at the extreme genome ends. In addition, abolishing the use of Le as a promoter for transcription enhanced replication. These analyses (i) identified signals at the termini required for transcription and replication and (ii) showed that Le functions as a less efficient promoter for replication than TrC at least in part because of its essential role in transcription. Consequently, these observations help explain the asymmetry of VSV replication which results in the synthesis of more negative- than positive-sense replication products in infected cells.
Whelan
S P
SP
Department of Microbiology, The Medical School, University of Alabama at Birmingham, Birmingham, Alabama, USA.
Wertz
G W
GW
D001483:Base Sequence; D016679:Genome, Viral; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic; D012367:RNA, Viral / Q000096:biosynthesis*; D011995:Recombination, Genetic; D014158:Transcription, Genetic; D014721:Vesicular stomatitis Indiana virus / Q000235:genetics*; D014779:Virus Replication
D012367:RNA, Viral