Fig. 2. Influence of histone H1 on transcription from the MMTV promoter. (A) In vitro transcription analysis of MMTV minichromosomes with increasing amounts of histone H1. Chromatin reconstituted with an increasing molar ratio of histone H1 was incubated with purified recombinant PR or/and NF1 as indicated and transcribed in vitro with HeLa nuclear extract. For each template, 25 ng of DNA were used in each reaction. Products were visualized by primer extension analysis and sequencing gel electrophoresis. The positions of the products from the wild-type MMTV promoter (wt) and from a control MMTV promoter lacking the HREs (–77) are indexed on the left. The synergism of PR and NF1 was calculated as: (activity PR + NF1)/(activity PR) + (activity NF1), and is indicated at the bottom. (B) Quantification of the wild-type MMTV transcripts. Transcription signals were quantified in a phosphoimager (Fuji) and are shown in relative units, with the transcription in the absence of H1 and activators (lane 1) set to 1. The numbers on the x-axis refer to the lanes in (A).