| Respiratory syncytial (RS) virus, the leading cause of bronchiolitis and pneumonia in infants, is a nonsegmented negative-sense RNA virus. The viral RNA polymerase accesses the genome at a single 3'-proximal promoter and transcribes the 10 genes sequentially. Each gene begins with a conserved gene start sequence, which directs initiation, and ends with a semiconserved gene end sequence, which directs polyadenylation and termination. The viral polymerase must terminate transcription of each gene to initiate transcription of the next gene downstream. The gene ends contain a semiconserved sequence, 3'-AUCAAUUNN-5', which is followed by a tract of U residues, thought to be the template for mRNA polyadenylation, and a nonconserved intergenic sequence.;We investigated the RNA sequences required for termination at the M gene end (3'-UUCAAUUAUUUUUU-5') in a dicistronic replicon. Individual nucleotide changes in the first nine positions of the M gene end reduced the efficiency of termination to varying levels (<10% to 85% of wild type termination), identifying sequences that were important for termination, but a U-to-A change at the first position increased termination. A U tract with at least four or five residues was required for efficient termination. The position of the U tract within the gene end was important for termination, as was the first nucleotide of the intergenic region downstream of a U4 tract.;The following consensus gene end sequence is proposed based on these data: 3'AUCAAUUN(A/U)UUUU(U/G/C)-5'. The 10 gene ends of RS virus have different sequences and termination efficiencies. The consensus gene end sequence predicted from our data helps explain how termination efficiency is modulated in RS virus. Our data also confirmed that termination can regulate transcription of individual genes.;In RS virus, transcription is also regulated by the trans-acting M2-1 protein, which reduces nonspecific termination within genes and may also decrease termination at gene ends. We investigated these effects of the M2-1 protein, and found that when polymerase terminated within a gene in the absence of M2-1 protein, it was able to initiate transcription of the downstream gene. The implications of these data for the mechanism of transcription and for gene regulation in RS virus are discussed. |
