6 research outputs found
Nucleosome Chiral Transition under Positive Torsional Stress in Single Chromatin Fibers
Using magnetic tweezers to investigate the mechanical response of single
chromatin fibers, we show that fibers submitted to large positive torsion
transiently trap positive turns, at a rate of one turn per nucleosome. A
comparison with the response of fibers of tetrasomes (the (H3-H4)2 tetramer
bound with ~50 bp of DNA) obtained by depletion of H2A-H2B dimers, suggests
that the trapping reflects a nucleosome chiral transition to a metastable form
built on the previously documented righthanded tetrasome. In view of its low
energy, <8 kT, we propose this transition is physiologically relevant and
serves to break the docking of the dimers on the tetramer which in the absence
of other factors exerts a strong block against elongation of transcription by
the main RNA polymerase.Comment: 33 pages (double spacing), 7 figure
Structural plasticity of single chromatin fibers revealed by torsional manipulation
Magnetic tweezers are used to study the mechanical response under torsion of
single nucleosome arrays reconstituted on tandem repeats of 5S positioning
sequences. Regular arrays are extremely resilient and can reversibly
accommodate a large amount of supercoiling without much change in length. This
behavior is quantitatively described by a molecular model of the chromatin 3-D
architecture. In this model, we assume the existence of a dynamic equilibrium
between three conformations of the nucleosome, which are determined by the
crossing status of the entry/exit DNAs (positive, null or negative). Torsional
strain, in displacing that equilibrium, extensively reorganizes the fiber
architecture. The model explains a number of long-standing topological
questions regarding DNA in chromatin, and may provide the ground to better
understand the dynamic binding of most chromatin-associated proteins.Comment: 18 pages, 7 figures, Supplementary information available at
http://www.nature.com/nsmb/journal/v13/n5/suppinfo/nsmb1087_S1.htm
Etude de la dynamique de la chromatine par des techniques de molécules uniques
PARIS7-Bibliothèque centrale (751132105) / SudocSudocFranceF
Compaction Kinetics on Single DNAs: Purified Nucleosome Reconstitution Systems versus Crude Extract
Kinetics of compaction on single DNA molecules are studied by fluorescence videomicroscopy in the presence of 1), Xenopus egg extracts and 2), purified nucleosome reconstitution systems using a combination of histones with either the histone chaperone Nucleosome Assembly Protein (NAP-1) or negatively charged macromolecules such as polyglutamic acid and RNA. The comparison shows that the compaction rates can differ by a factor of up to 1000 for the same amount of histones, depending on the system used and on the presence of histone tails, which can be subjected to post-translational modifications. Reactions with purified reconstitution systems follow a slow and sequential mechanism, compatible with the deposition of one (H3-H4)(2) tetramer followed by two (H2A-H2B) dimers. Addition of the histone chaperone NAP-1 increases both the rate of the reaction and the packing ratio of the final product. These stimulatory effects cannot be obtained with polyglutamic acid or RNA, suggesting that yNAP-1 impact on the reaction cannot simply be explained in terms of charge screening. Faster compaction kinetics and higher packing ratios are reproducibly reached with extracts, indicating a role of additional components present in this system. Data are discussed and models proposed to account for the kinetics obtained in our single-molecule assay