Specific transcription of an adenoviral gene that possesses no TATA sequence homology in extracts of HeLa cells

Transcription of the adenovirus type 2 (Ad2) IVa₂ gene, which contains no TATA-like sequence in the region immediately upstream of the IVa₂ cap sites, has been examined in extracts of HeLa cells. Run-off transcripts of the predicted length of those initiated at the IVa₂ cap sites were synthesized from different Ad2 DNA templates, each of which also contained the major late transcriptional control region. Mapping of the 5' ends of the RNA made from one template by a nuclease protection assay established the fidelity of initiation of IVa₂ transcription in vitro. The efficiency of IVa₂ expression in whole HeLa extracts was influenced quite dramatically by monovalent and divalent metal ion concentrations and the concentration of extract protein present in the reaction mixture. Under certain conditions, IVa₂ run-off transcripts were made almost as efficiently as those from the Ad2 major late transcriptional control region. However, conditions promoting optimal IVa₂ transcription in vitro did not favor recognition of the major late transcriptional control region, and vice versa: the synthesis of IVa₂ and major late run-off transcripts responded differently to all parameters tested.