Results: 4

1.
Figure 3

Figure 3. From: Histone Sin mutations promote nucleosome traversal and histone displacement by RNA polymerase II.

Advancing RNA polymerase 41 bp into a Sin mutant nucleosome results in nuclease accessibility of downstream DNA. (A) DNase I footprinting of non-transcribed 603 nucleosomes (left gel) or EC+41–603 nucleosome complexes (right gel). Histones were wild type (W) or H3 T118I (S); D lanes are free DNA controls. The dotted line indicates protection by the EC+41 complex. (B) Scans of the footprints in (A): naked DNA (black), wild-type (upper, red) or Sin (lower, red) nucleosomes, and corresponding EC+41 complexes (green). EC, elongation complex; H3, histone H3; M, size markers; N, nucleosome; RNAP, RNA polymerase; wt, wild type.

Fu-Kai Hsieh, et al. EMBO Rep. 2010 September;11(9):705-710.
2.
Figure 1

Figure 1. From: Histone Sin mutations promote nucleosome traversal and histone displacement by RNA polymerase II.

Nucleosomes containing Sin mutant histones or lacking amino-terminal tails provide a reduced barrier to traversal by polymerase II. (A) Nucleosomes were assembled on the 603 template using wild-type (n/x) or tailless (g/x) H2A/H2B, and wild-type (x/n) or Sin mutant H3 and H4. RNA in yeast polymerase II complexes was initially pulse-labelled and then extended at the indicated KCl concentrations with excess unlabelled NTPs. The locations of the nucleosome (oval), the nucleosome dyad (square), the main pol II pause sites and the position of the run-off transcript are shown. (B) As in (A), except that human pol II was used. KCl concentrations, sarkosyl addition (1%) and no-chase controls (nc) are indicated. (C) As in (B), except that the template was 603R. g, globular; H3, histone H3; H4, histone H4; n, native; pol II, polymerase II.

Fu-Kai Hsieh, et al. EMBO Rep. 2010 September;11(9):705-710.
3.
Figure 2

Figure 2. From: Histone Sin mutations promote nucleosome traversal and histone displacement by RNA polymerase II.

Nucleosomes containing Sin mutant histones are more likely to dissociate during pol II transcription. (A) DNA-labelled nucleosomes containing wild-type or Sin mutant histones were transcribed by yeast pol II at the indicated KCl concentrations; intact complexes were then resolved from free DNA by native polyacrylamide gel electrophoresis. Only the region of the gel in which free DNA appears is shown; an example of the entire native gel from such an experiment is shown in supplementary Fig S4 online. No-chase control (nc): Transcription was performed in the absence of UTP and pol II was stalled upstream of the nucleosome. (B) The amounts of histone-free DNA produced after transcription at 150 mM KCl were quantified and normalized to the overall amount of active ECs. EC, elongation complex; H3, histone H3; H4, histone H4; pol II, polymerase II.

Fu-Kai Hsieh, et al. EMBO Rep. 2010 September;11(9):705-710.
4.
Figure 4

Figure 4. From: Histone Sin mutations promote nucleosome traversal and histone displacement by RNA polymerase II.

Proposed mechanism of transcription through nucleosomes. Pol II enters the nucleosome (A), partly displaces upstream DNA (B) and forms a zero (∅)-loop at +39 (C), inducing reversible uncoiling of downstream DNA (D). On wild-type nucleosomes, continued transcription is accompanied by nucleosome recovery. However, strong DNA–histone interactions (blue squares) are weakened in Sin nucleosomes (purple squares), causing a larger downstream DNA region to be displaced (E), favouring nucleosome displacement. Inset panel, from Luger et al (1997): DNA, grey; H3, blue; H4, green; H3 residues Arg 116 and Thr 118, yellow; H4 residues Val 43 and Arg 45, orange; histone–DNA contacts disrupted by Sin mutants (Muthurajan et al, 2004), red. The white diamond indicates the nucleosome dyad. H3, histone H3; H4, histone H4; pol II, polymerase II.

Fu-Kai Hsieh, et al. EMBO Rep. 2010 September;11(9):705-710.

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