Transcription of bacteriophage M13 DNA: existence of promoters directly preceding genes III, VI, and I.

In vitro transcription and coupled transcription-translation studies have been performed with restriction fragments of bacteriophage M13 replicative-form DNA which contain either gene III, gene VI, or gene I. It could be demonstrated that DNA fragments which contain gene III were able to direct the synthesis of gene III protein. Fragments which encompassed genes VI and I gave rise to the synthesis of gene I protein only, whereas gene I-containing fragments were able to direct the synthesis of gene I protein. None of the fragments studied gave rise to a detectable level of gene VI protein, although an RNA transcript of gene VI could readily be obtained during in vitro transcription of the relevant gene VI-containing DNA fragments. From these results we have concluded that the promoters A0.44 and A0.49 are located in front of genes VI and I, respectively, and that gene III is also equipped with a promoter (X0.25). Introduction of a single cleavage within the gene III region does not abolish the expression of genes VI and I in vitro. Hence, the expression of these genes is not solely dependent on the initiation of RNA synthesis at the gene III promoter or on leakage of transcription through the central termination site (T0.25), but is also determined by the initiation frequency of RNA synthesis at their individual promoters.