TY - JOUR
T1 - The nature of the nucleosomal barrier to transcription
T2 - Direct observation of paused intermediates by electron cryomicroscopy
AU - Bednar, Jan
AU - Studitsky, Vasily M.
AU - Grigoryev, Sergei A.
AU - Felsenfeld, Gary
AU - Woodcock, Christopher L.
N1 - Funding Information:
We wish to thank R. Horowitz who helped at the earlier stage of the work, Dr. O. Kulaeva for help with plasmid construction, and Dr. L. Lutter, Dr. C. Martin, and Dr. D. Clark for valuable discussions and comments on the manuscript. The work was supported in part by National Institutes of Health grants GM51352 to S. A. G., GM58650 to V. M. S., and GM43786 to C. L. W. The Central Microscopy Facility at the University of Massachusetts–Amherst is supported in part by National Science Foundation grant BIR-9419676.
PY - 1999/9
Y1 - 1999/9
N2 - Transcribing SP6 RNA polymerase was arrested at unique positions in the nucleosome core, and the complexes were analyzed using biochemical methods and electron cryomicroscopy. As the polymerase enters the nucleosome, it disrupts DNA-histone interactions behind and up to ~20 bp ahead of the elongation complex. After the polymerase proceeds 30-40 bp into the nucleosome, two intermediates are observed. In one, only the DNA ahead of the polymerase reassociates with the octamer. In the other, DNA both ahead of and behind the enzyme reassociates. These intermediates present a barrier to elongation. When the polymerase approaches the nucleosome dyad, it displaces the octamer, which is transferred to promoter-proximal DNA.
AB - Transcribing SP6 RNA polymerase was arrested at unique positions in the nucleosome core, and the complexes were analyzed using biochemical methods and electron cryomicroscopy. As the polymerase enters the nucleosome, it disrupts DNA-histone interactions behind and up to ~20 bp ahead of the elongation complex. After the polymerase proceeds 30-40 bp into the nucleosome, two intermediates are observed. In one, only the DNA ahead of the polymerase reassociates with the octamer. In the other, DNA both ahead of and behind the enzyme reassociates. These intermediates present a barrier to elongation. When the polymerase approaches the nucleosome dyad, it displaces the octamer, which is transferred to promoter-proximal DNA.
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U2 - 10.1016/S1097-2765(00)80339-1
DO - 10.1016/S1097-2765(00)80339-1
M3 - Article
C2 - 10518218
AN - SCOPUS:0033197561
SN - 1097-2765
VL - 4
SP - 377
EP - 386
JO - Molecular cell
JF - Molecular cell
IS - 3
ER -