Sheinin MY et al. (2013), Torque modulates nucleosome stability and facil...

XB-ART-48968

Nat Commun 2013 Jan 01;4:2579. doi: 10.1038/ncomms3579.

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Torque modulates nucleosome stability and facilitates H2A/H2B dimer loss.

Sheinin MY , Li M , Soltani M , Luger K , Wang MD .

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The nucleosome, the fundamental packing unit of chromatin, has a distinct chirality: 147 bp of DNA are wrapped around the core histones in a left-handed, negative superhelix. It has been suggested that this chirality has functional significance, particularly in the context of the cellular processes that generate DNA supercoiling, such as transcription and replication. However, the impact of torsion on nucleosome structure and stability is largely unknown. Here we perform a detailed investigation of single nucleosome behaviour on the high-affinity 601-positioning sequence under tension and torque using the angular optical trapping technique. We find that torque has only a moderate effect on nucleosome unwrapping. In contrast, we observe a dramatic loss of H2A/H2B dimers on nucleosome disruption under positive torque, whereas (H3/H4)₂ tetramers are efficiently retained irrespective of torsion. These data indicate that torque could regulate histone exchange during transcription and replication.

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Species referenced: Xenopus laevis
Genes referenced: h2ac21 h2bc21

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	Fig. 2. Characterizing nucleosome disruption under force and torque(a) Sudden release of the inner (filled squares) and outer (open circles) turns of DNA in a nucleosome. Error bars are SEM. (b) Mean disruption forces of the inner (filled squares) and outer (open circles) turns of DNA in a nucleosome. Error bars are SEM. Between 25 and 64replicates have been obtained for each torque (Table 1).(c) Fraction of traces that displayed a sudden outer turn release as a function of torque. Error bars are standard errors of the binomial distribution (see Methods).
	Fig. 3. Tetrasome force-extension curvesNaked DNA (grey), recombinant tetrasome (red), recombinant nucleosome with a sudden lower force disruption (blue), all stretched under no torsion.
	Fig. 4. Characterizing tetrasome disruption under force and torque(a) Sudden release of the inner (filled squares) and outer (open circles) turns of DNA in a tetrasome and a nucleosome. Error bars are SEM. (b) Mean disruption forces of the inner (filled squares) and outer (open circles) turns of DNA in a tetrasome and a nucleosome. Error bars are SEM. Between 15 and 37replicates have been obtained for each torque (Table 1).(c) Fraction of traces that displayed a sudden outer turn release as a function of torque. Red color refers to Xenopus recombinant tetrasome, blue - Xenopus recombinant nucleosome, black â HeLa purified nucleosome. Error bars are standard errors of the binomial distribution (see Methods).
	Fig. 5. Histone fate upon disruption under torque(a) Experimental strategy to examine the nucleosome stability under torque. A nucleosomal DNA molecule was supercoiled to a desired torsion, stretched at 2 pN/s from 1 pNup to 30 pNof force to disrupt the nucleosome before being relaxed over 1 s to 0.7 pN. It was then wound to +38 pNÂ·nmand stretched again to determine which histones remained on the DNA. The time between relaxation and the second stretching was always kept at 30 s. (b) The probability of observing a sudden disruption at low force during the second stretch (right inset) was normalized to the corresponding value of the control stretch at +38 pNÂ·nm of torque. Normalized probability of observing a sudden low force disruption is equivalent to the probability of retaining H2A/H2B after the initial stretch. Between 21 and 28replicates have been obtained for each torque value (Table 1). Error bars are standard errors of the ratio of two binomial distributions (see Methods).(c) The probability of observing a sudden high force disruption (right inset) signifies the probability of retaining (H3/H4)2 after the initial stretch. Between 21 and 28replicates have been obtained for each torque (Table 1) Insets show a representative trace of the force-extension curve resulting from a second stretch. Error bars are standard errors of the binomial distribution (see Methods).(d) Cartoons of predominant histones remaining on the DNA after disruption.
	Fig. 6. A cartoon illustrating RNA polymerase II elongation through chromatin. During transcription under low torsion, nucleosomes remain largely intact after transcription. In contrast, during transcription under high torsion, significant dimer loss occurs in nucleosome after transcription. (H3/H4)2 tetramer is shown in brown, and H2A/H2B dimer in orange.

References [+] :

Alabert, Chromatin replication and epigenome maintenance. 2012, Pubmed