Analysis of the role of the transcription factor ATF in the assembly of a functional preinitiation complex.

In the accompanying paper (Horikoshi et al., 1988) the interaction between ATF and the general transcription initiation factors was analyzed by DNAase I footprinting experiments. Here, we use transcription assays to investigate the role of ATF in the assembly of a functional preinitiation complex. Addition of an oligonucleotide containing an ATF binding site inhibits E4 transcription by sequestering ATF. However, following preincubation of the E4 promoter in the nuclear extract, transcription is refractory to inhibition by the ATF oligonucleotide. Formation of this oligonucleotide-refractory complex occurs at an early stage in the overall transcription initiation reaction and is dependent upon ATF and the general transcription factors RNA polymerase II, TFIIB, and TFIID. This latter result suggests that the assembly and maintenance of a functional preinitiation complex involves cooperative interactions among the various transcription factors. The general transcription factor TFIIE, although required for transcriptional activity, is not involved in the assembly of an ATF oligonucleotide-refractory complex. Our results support the possibility that ATF may be required only transiently for assembly of a functional preinitiation complex.