ASSOCIATION OF NUCLEOSOME-FREE REGIONS AND BASAL TRANSCRIPTION FACTORS WITH IN-VIVO-ASSEMBLED CHROMATIN TEMPLATES ACTIVE IN-VITRO

Using SV40 minichromosomes assembled in vivo, we have studied the rela tionship between a nucleosome-free promoter-region and initiation of t ranscription by RNA polymerase II on chromatin templates in vitro. Our data suggest that accessibility of DNA to transcription factors, prog rammed into the structure of the chromatin, is crucial for initiation of transcription. First, minichromosomes competent to be transcribed i n vitro contained nucleosome-free promoter regions. Second, tsC219 min ichromosomes, most of which contain the nucleosome-free promoter regio n, supported transcription more efficiently both in vivo and in vitro than wild-type minichromosomes, in which only a subset contain the nuc leosome-free region. We have also identified basal transcription facto rs associated with the in vivo-assembled chromatin templates. A striki ng correlation was observed between minichromosomes associated with in vivo initiated RNA polymerases and those associated with the basal tr anscription factors TFIID and TFIIE/F, and to a lesser extent, TFIIB. Of these associated factors, only TFIID was poised for ready assembly into preinitiation complexes and therefore for subsequent initiation o f transcription. However, an active chromatin template could also be m aintained in the absence of the binding of TFIID. Finally, our data ar e consistent with the presence of TFIIF in elongating ternary complexe s on the chromatin templates.