Bacteriophage N4 is lytic phage specific for Escherichia soli K-12. Transcription of N4 genome can be divided into three stages, early, middle, and late, depending on the time of appeareance of the transcripts and the RNA polymerase species which catalyze their synthesis.
N4 is unique among DNA-containing bacteriophages because transcription of the early region of the genome is carried out by its own coded RNA polymerase, called the virion encapsulated RNA polymerase. This polymerase is present in the virion and injencted along with the genome upon infection.
Early transcription requirs active E. coli DNA gyrase and single-stranded DNA binding protein (SSB). The N4 virion RNA polymerase promoters comprise conserved sequence present at -18 to + 1, and a set of inverted repeats centered at -12. These sequences are recognized on a single-stranded template or on a double-stranded templaten only if the DNA is supercoiled and SSB is present. Activation of transcription on supercoiled template is SSB specific. Mutational analysis of the promoters has reveal-
that both recognition of conserved sequences and DNA secondary structure at the promoter are required for pro ductive virion RNA polymerase-promoter interaction.
Three products of N4 early transcription, p4, p7, and p17, are required for the synthesis of middle transcription. Two of them, p4 and p7, compose N4 RNA polymerase II which is responsible for middle transcription. This polymerase requires a third protein, p17, for specific promoter recognition. Middle promoters have been determined by Sl-nuclease mapping. It has two sets of con-served, sequences at variable distances.
Late transcription requires the activity of the E. coli o 70-holoenzyme. In vivo sites of transcription initiation have been determined. These sites show only 50% homology to the E. coli o70-consensus sequence and no interhomology at the -10 or 35 region. Late transcription is not dependent on phage DNA replication but does require active N4 coded, single-stranded DNA binding protein (N4SSB). In vitro, N4 late transcription initiation sites are utilized poorly by purified E. coli a70-holoenzyme.
N4SSB allows E. coli 670-holoenzyme to utilize these sites efficiently. The E. coli o70-holoenzyme carries out transcription at a late promoter efficiently without any aid of N4SSB when a late promoter placed in a plasmid. It has been shown that strong activity of a late promoter placed in a plasmid is due to the negative superhelicity. Since supercolling of the template circumvents the requirement of N4SSB for efficient transcription, both N4SSB and supercoiling may play a similar role in the initiation step.
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