Highly diverse intergenic regions of the paramyxovirus simian virus 5 cooperate with the gene end U tract in viral transcription termination and can influence reinitiation at a downstream gene

A dicistronic minigenome containing the M-F gene junction was used to deter mine the role of the simian virus 5 (SV5) intergenic regions in transcripti on, The M-F junction differs from the other SV5 junctions by having a short M gene end U tract of only four residues (U4 tract) and a 22-base M-F inte rgenic sequence between the M gene end and F gene start site. Replacing the 22-base M-I: intergenic region with nonviral sequences resulted in a minig enome template (Rep 22) that was defective in termination at the end of the hi gene, Efficient M gene termination could be restored to the mutant Rep 22 template in either of two ways: by increasing the U tract length from fo ur to six residues or by restoring a G residue immediately downstream of th e wild-type (WT) U4 tract. In a dicistronic SH-HN minigenome, a U4-G combin ation was functionally equivalent to the naturally occurring SH U6-A gene e nd in directing SD transcription termination. In addition to affecting term ination, the M-F intergenic region also influenced polymerase reinitiation, In the context of the WT U4-G IM gene end, substituting nonviral sequences into the M-F intergenic region had a differential effect on F gene reiniti ation, where some but not all nonviral sequences inhibited reinitiation, Th e inhibition of F gene reinitiation correlated crith foreign sequences havi ng a high C content, Deleting 6 bases or inserting 18 additional nucleotide s into the middle of the 22-base M-F intergenic segment did not influence M gene termination or F gene reinitiation, indicating that M-F intergenic le ngth per se is not a important factor modulating the SV5 polymerase activit y. Our results suggest that the sequence diversity at an SV5 gene junction reflects specific combinations which may differentially affect SV5 gene exp ression and provide an additional level of transcriptional control beyond t hat which results from the distance of a gene from the 3' end promoter.