FIG. 1.

(A) The reporter DNA construct pMMTV:M13 used for injection. Solid black arrows, primers used for primer extension analysis of SacI in situ accessibility and DMS methylation protection. The restriction enzyme sites shown are referred to in the text. Arrow (+1), transcription start site. GRE I to IV (white boxes), the NF1 binding site (light gray), Oct1-binding sites (black), and the TATA box (dark gray) and their indicated cognate DNA binding segments are displayed with the same shading. Black dots, protected guanines in DMS in vivo footprinting. (B) Autoradiography of Xenopus oocyte extract 24 h after injection of 6 ng of GR mRNA alone or together with 12 ng of NF1 mRNA followed by incubation in [³⁵S]methionine (see Materials and Methods). (C) DMS methylation protection analysis of the MMTV DNA segment −200 to −50 for oocytes injected with 3 ng of pMMTV:M13 DNA and GR-NF1 mRNA as for panel B and the next day not treated (−) or treated (+) with 1 μM synthetic glucocorticoid TA. Radioactivity scans show the two lanes with highlighted hormone-dependent effects: methylation-protected bands (white circles) and a hypermethylated band (black dot).