FIG. 7.
pX recovers TFIIB activity of the C34,37S mutant that it binds. (A) Increasing amounts of the TFIIB mutants were cotransfected with the Gal4-p53 and the Gal4-luciferase reporter, with or without pX expressor as indicated. Luciferase activity in the presence of pX relative to activity in the absence of pX is presented as fold pX effect. Cytoplasmic fractions of cells transfected as indicated were analyzed by luciferase assay. Two different pX mutants, Δ67-86 and Δ105-154, were used. (B) DNA templates bearing the U-less cassette of 112 bases, under the regulation of adenovirus type 5 major late promoter TATA and UASG elements, were used for in vitro-reconstituted transcription reactions. All reactions contained the DEA and DEB fractions, as described in reference 23. Two human TFIIB proteins (30 ng of either wt [hTFIIB] or C34,37S [hTFIIBm]), 10 ng of Gal4-VP16 protein, and 10 ng of recombinant pX were added to the reactions where indicated. Transcription products were purified and analyzed by denaturing gel electrophoresis.