Dataset Information

Nucleosome hopping and sliding kinetics determined from dynamics of single chromatin fibers in Xenopus egg extracts.

ABSTRACT: Chromatin function in vivo is intimately connected with changes in its structure: a prime example is occlusion or exposure of regulatory sequences via repositioning of nucleosomes. Cell extracts used in concert with single-DNA micromanipulation can control and monitor these dynamics under in vivo-like conditions. We analyze a theory of the assembly-disassembly dynamics of chromatin fiber in such experiments, including effects of lateral nucleosome diffusion ("sliding") and sequence positioning. Experimental data determine the force-dependent on- and off-rates as well as the nucleosome sliding diffusion rate. The resulting theory simply explains the very different nucleosome displacement kinetics observed in constant-force and constant-pulling velocity experiments. We also show that few-piconewton tensions comparable to those generated by polymerases and helicases drastically affect nucleosome positions in a sequence-dependent manner and that there is a long-lived structural "memory" of force-driven nucleosome rearrangement events.

SUBMITTER: Ranjith P

PROVIDER: S-EPMC1959436 | biostudies-literature | 2007 Aug

REPOSITORIES: biostudies-literature

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Nucleosome hopping and sliding kinetics determined from dynamics of single chromatin fibers in Xenopus egg extracts.

Ranjith Padinhateeri P Yan Jie J Marko John F JF

Proceedings of the National Academy of Sciences of the United States of America 20070814 34

Chromatin function in vivo is intimately connected with changes in its structure: a prime example is occlusion or exposure of regulatory sequences via repositioning of nucleosomes. Cell extracts used in concert with single-DNA micromanipulation can control and monitor these dynamics under in vivo-like conditions. We analyze a theory of the assembly-disassembly dynamics of chromatin fiber in such experiments, including effects of lateral nucleosome diffusion ("sliding") and sequence positioning. ...[more]

PMID: 17698962

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Project description:BackgroundLinker histone H1 has been studied in vivo and using reconstituted chromatin, but there have been few systematic studies of the effects of the cellular environment on its function. Due to the presence of many other chromatin factors and specific chaperones such as RanBP7/importin beta that regulate histone H1, linker histones likely function differently in vivo than in purified systems.Methodology/principal findingsWe have directly compared H1 binding to sperm nuclei in buffer versus Xenopus egg extract cytoplasm, and monitored the effects of adding nuclear import chaperones. In buffer, RanBP7 decondenses sperm nuclei, while H1 binds tightly to the chromatin and rescues RanBP7-mediated decondensation. H1 binding is reduced in cytoplasm, and H1 exhibits rapid FRAP dynamics in cytoplasm but not in buffer. RanBP7 decreases H1 binding to chromatin in both buffer and extract but does not significantly affect H1 dynamics in either condition. Importin beta has a lesser effect than RanBP7 on sperm chromatin decondensation and H1 binding, while a combination of RanBP7/importin beta is no more effective than RanBP7 alone. In extracts supplemented with RanBP7, H1 localizes to chromosomal foci, which increase after DNA damage. Unlike somatic H1, the embryonic linker histone H1M binds equally well to chromatin in cytoplasm compared to buffer. Amino-globular and carboxyl terminal domains of H1M bind chromatin comparably to the full-length protein in buffer, but are inhibited ?10-fold in cytoplasm. High levels of H1 or its truncations distort mitotic chromosomes and block their segregation during anaphase.Conclusion/significanceRanBP7 can decondense sperm nuclei and decrease H1 binding, but the rapid dynamics of H1 on chromatin depend on other cytoplasmic factors. Cytoplasm greatly impairs the activity of individual H1 domains, and only the full-length protein can condense chromatin properly. Our findings begin to bridge the gap between purified and in vivo chromatin systems.

Dataset Information

Nucleosome hopping and sliding kinetics determined from dynamics of single chromatin fibers in Xenopus egg extracts.

Publications

Nucleosome hopping and sliding kinetics determined from dynamics of single chromatin fibers in Xenopus egg extracts.

Similar Datasets

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