Eukaryotic ternary transcription complexes. II. An approach to the determination of chromatin conformation at the site of transcription.

AUTOR(ES)

Sargan, D R

RESUMO

Digestion of rat liver nuclei by endogenous nucleases or micrococcal nuclease releases a chromatin fraction containing RNA polymerases I and II bound to DNA fragments in ternary transcription complexes. To label the DNA in these transcription complexes, the polymerases were allowed to add radioactively labelled ribonucleotides in vitro to in vivo-initiated RNA chains. During this transcription step, nucleic acids were photochemically cross-linked using 8-methoxypsoralen. Nucleic acids in transcription complexes were then sized by gel electrophoresis. Under conditions where RNA polymerases I and II were active in vitro, most of the labelled DNA was found in a series of fragments of sizes which were multiples of approximately 200 base-pairs. When polymerase I alone was active, the smallest member of this series carried the bulk of the label; when polymerase II also was active, a significant proportion of the label was carried on the dimer and higher oligomers. Proteins other than polymerase alone are shown to be responsible for the pattern of DNA fragments protected from nucleases. Therefore active RNA polymerases I and II in vivo are in close proximity to structures protecting DNA fragments, the sizes of which are similar to those found in nucleosomes. We have yet to establish that these structures are composed of histones.

Documentos Relacionados

• Eukaryotic ternary transcription complexes. I. The release of ternary transcription complexes of RNA polymerases I and II by the endogenous nucleases of rat liver nuclei.
• The active site of RNA polymerase II participates in transcript cleavage within arrested ternary complexes.
• The sv40 transcription complex. II. Non-dissociation of protein from SV40 chromatin during transcription.
• Eukaryotic ternary transcription complexes: transcription complexes of RNA polymerase II are associated with histone-containing, nucleosome-like particles in vivo.
• Incorporation of chromosomal proteins HMG-14/HMG-17 into nascent nucleosomes induces an extended chromatin conformation and enhances the utilization of active transcription complexes.