Nucleic Acids Research, 1993, Vol. 21, No. 15 3459-3468
© 1993
MOLECULAR BIOLOGY |
Association of nucleosome-free regions and basal transcription factors with in vivo-assembled chromatin templates active in vitro
Laboratory of Eukaryotic Transcription, Dana-Farber Cancer Institute and Department of Microbiology and Molecular Genetics, Harvard Medical School Boston, MA 02115, USA
*To whom correspondence should be addressed
Received April 22, 1993. Revised June 15, 1993. Accepted June 15, 1993.
Using SV40 minichromosomes assembled in vivo, we have studied the relationship between a nucleosome-free promoter-region and initiation of transcription by RNA polymerase II on chromatin templates in vitro. Our data suggest that accessibility of DNA to transcription factors, programmed into the structure of the chromatin, is crucial for initiation of transcription. First, minichromosomes competent to be transcribed in vitro contained nucleosome-free promoter regions. Second, tsC219 minichromosomes, most of which contain the nucleosome-free promoter region, supported transcription more efficiently both in vivo and in vitro than wild-type minichromosomes, in which only a subset contain the nucleosome-free region. We have also identified basal transcription factors associated with the in vivo-assembled chromatin templates. A striking correlation was observed between minichromosomes associated with in vivo initiated RNA polymerases and those associated with the basal transcription factors TFIID and TFIIE/F, and to a lesser extent, TFIIB. Of these associated factors, only TFIID was polsed for ready assembly into preinitiation complexes and therefore for subsequent initiation of transcription. However, an active chromatin template could also be maintained in the absence of the binding of TFIID. Finally, our data are consistent with the presence of TFIIF in elongating ternary complexes on the chromatin templates.
+Present address: Department of Psychiatry, University of Iowa College of Medicine, Iowa City, LA 52242, USA
Present address: Department of Molecular Biology, Institut Pasteur, 75724 Paris, France
¶Present address: Burroughs Wellcome Company, Research TrianglePark, NC 27709, USA
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