Positive and negative control sequences within the distal domain of the adenovirus IVa2 promoter overlap with the major late promoter.

The RNA initiation sites of the adenovirus IVa2 and major late promoters (MLP) are separated by 210 base pairs and transcribed from opposite DNA strands. We had previously shown that they contained overlapping promoter sequences (V. Natarajan, M. J. Madden, and N. P. Salzman, Proc. Natl. Acad. Sci. U.S.A. 81:6290-6294, 1984). The transcription efficiencies of these two promoters were studied in vitro with templates of covalently closed circular DNAs that contained various deletion and point mutants. The distal control region of the IVa2 promoter that is located at nucleotide position (np) -152 to -242 from the RNA initiation site consists of at least two domains. The first distal domain, present between np -152 and -179, is necessary for efficient transcription of the IVa2 promoter, and it overlaps with sequences that have been shown to be necessary for efficient transcription of MLP. This region may serve as the entry site for the transcription machinery. The second distal domain consists of sequences present between np -211 and -242. These sequences are contained at the 5' end in the MLP transcript, and they inhibit transcription from the IVa2 promoter. However, these sequences are not necessary for transcription of the MLP with a covalently closed template but are needed for transcription with a linear template. The TATA box that is located at np -180 to -186 between these two domains has a critical role for efficient transcription of the MLP. A point mutation that reduces transcription from MLP by more than 80% stimulates transcription from IVa2 promoter by 10-fold. This finding is consistent with the proposal that MLP and IVa2 promoters share an entry site for transcription machinery and compete for its use.