1887

Abstract

Paramyxovirus genomes contain a linear array of five to ten genes sequentially transcribed by the viral RNA polymerase. mRNA synthesis initiates at a nucleotide signal () within the genomic promoter located at the genome 3′ end. To gain information about the mechanism involved in transcription initiation, a search was carried out for upstream nucleotides required for and the effects of the nucleocapsid protein (N) phase context on transcription regulation were determined. For both purposes, tandem promoter mini-genomes carrying a transcription signal ectopically positioned downstream of a replication-only signal were used. The requirement for hygromycin resistance gene expression was used in an attempt to select essential nucleotides within randomized stretches of nucleotides. Nucleotide insertions or deletions were also made on either side of the transcription signal to change its original N phase context in the five remaining possibilities and GFP expression from these modified signals was assessed. Cell cultures resistant to hygromycin treatment were readily obtained following amplification of mini-genomes harbouring randomized sequences. However, selected nucleotides upstream of could not be identified under conditions where nucleotides within were selected. In contrast, it was observed that changing the N phase context progressively decreased transcription by five- to tenfold. These results are discussed in relation to two different mechanisms of transcription initiation.

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2007-05-01
2019-11-13
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