Fig. 4.

(A) Shown is a schematic of parallel in vitro transcription reactions using pG5MLT reconstituted chromatin as template in which two levels of E1A 1–80 (62.5 and 125 ng) were added at various stages. (B) Shown are transcripts from the reactions depicted above. The relative amounts of transcription are indicated. At pre-activation, strong, dose-dependent repression of transcription from the pG5MLT chromatin template by E1A 1–80 is seen, an average of 49% of the control activity for 62.5 ng and 12% for 125 ng of E1A 1–80 with a standard deviation (SD) of 6% and 9% respectively from 8 experiments. At activation, strong dose-dependent repression is also seen, an average of 56% of control for 62.5 ng and 22% for 125 ng of E1A 1–80 with a SD of 8% and 9% respectively from 8 experiments. At PIC formation, significantly enhanced activity is seen when of E1A 1–80 is added; 277% of control activity for 62.5 ng and 148% for 125 ng with a SD of 17% and 19% respectively from 5 experiments. At transcription-initiation, slightly enhanced activity is again seen when 62.5 ng of E1A 1–80 is added; an average of 116% control activity with a SD of 27% from 5 experiments. However, when 125 ng of E1A 1–80 is added, significant repression is seen with an average of 37% of the control activity with a SD of 16% for 5 experiments. At early transcription, strong dose-dependent repression is again seen, 72% of control activity for 62.5 ng and 41% with 125 ng of E1A 1–80 with an SD of 6% and 9% respectively from 8 experiments. (C) Shown is a parallel temporal analysis using pG5MLT as naked DNA template. In all stages, E1A 1–80 represses transcription in a dose-dependent manner.