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Nucleic Acids Research, 1984, Vol. 12, No. 19 7423-7433
© 1984

MOLECULAR BIOLOGY

The adenovirus major late promoter TATA box and initiation site are both necessary for transcription in vitro

Michael F. Concino+, Richard F. Lee, James P. Merryweather* and Roberto Weinmann

The Wistar Institute of Anatomy and Biology 36th Street at Spruce, Philadelphia, PA 19104 *Chiron Corporation Emeryville, CA 94608, USA

Received May 29, 1984. Revised September 13, 1984. Accepted September 13, 1984.

Mutagenized DNA templates and HeLa whole cell extracts were used to study the effects of promoter-specific base changes on in vitro transcription. DNA templates where the initiating adenine (+1) was changed to thymidine (AT+1) 1n the adenovirus 2 major late transcription unit were transcribed with 50% efficiency of the unaltered template. We have described a mutant at the TATA box, where the A at position –28 was changed to a C (AC-28). Transcription efficiency was reduced to less than 20% of control in the AC-28 mutant (Concino et al., 1983, J. Biol. Chem. 258: 8493–8496). Primer extension analysis revealed increased 5' end heterogeneity for in vitro transcripts derived from AC-28 and AT+1 DNA templates. Specific transcription was completely abolished from AT+1 DNA templates when a second change was introduced within the TATA sequence, creating a double mutant (AC-28 . AT+1). Neither the AC-28 nor the AT+1 change alone had such an effect, suggesting a coordinated interaction in transcription initiation involving both the TATA box and the Initiation site.

+Present address: Damon Biotech, 119 FOurth Avenue, needham Heights, MA 02194, USA


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