In cells productively infected with adenovirus type 5, transcription is not 
terminated between the E1a gene and the adjacent downstream E1b gene. Insertion 
of the mouse beta(maj)-globin transcription termination sequence (GGT) into the 
E1a coding region dramatically reduces early, but not late, E1b expression (E. 
Falck-Pedersen, J. Logan, T. Shenk, and J. E. Darnell, Jr., Cell 40:897-905, 
1985). In the study described herein, we showed that base substitution mutations 
in the globin DNA that specifically relieved transcription termination also 
restored early E1b promoter activity in cis, establishing that maximal early E1b 
expression requires readthrough transcription originating from the adjacent 
upstream gene. To identify potential targets of readthrough activation, a series 
of recombinant viruses with double mutations was constructed. Each double-mutant 
virus strain had the transcription termination sequences in the first exon of 
E1a and a deletion within the transcription control region of E1b. Early E1b 
expression from the double-mutant strains was more defective than that from 
strains containing either mutation alone, indicating that the deleted regions 
(positions -362 to -35) are not the target for readthrough activation. Two 
findings suggested that a cis-dominant property of early viral templates is 
important for readthrough activation. First, the early E1b defect caused by the 
GGT insertion was not complemented in trans by factors present in late-infected 
cells. Second, restoration of E1b transcription at late times occurred 
concurrently with viral DNA replication. Readthrough activation may help convert 
virion DNA into a transcriptionally competent template prior to DNA replication 
and late transcription.