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PMID: 11425866 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't

Kinetics of core histones in living human cells: little exchange of H3 and H4 and some rapid exchange of H2B.

The Journal of cell biology ·Vol. 153 ·No. 7 ·2001-06-25 ·Pages 1341-53

Kimura H, Cook PR

Abstract

Histones H2A and H2B form part of the same nucleosomal structure as H3 and H4. Stable HeLa cell lines expressing histones H2B, H3, and H4 tagged with green fluorescent protein (GFP) were established; the tagged molecules were assembled into nucleosomes. Although H2B-GFP was distributed like DNA, H3-GFP and H4-GFP were concentrated in euchromatin during interphase and in R-bands in mitotic chromosomes. These differences probably result from an unregulated production of tagged histones and differences in exchange. In both single cells and heterokaryons, photobleaching revealed that H2B-GFP exchanged more rapidly than H3-GFP and H4-GFP. About 3% of H2B exchanged within minutes, whereas approximately 40% did so slowly (t(1/2) approximately 130 min). The rapidly exchanging fraction disappeared in 5,6-dichloro-1-beta-d-ribofuranosylbenzimidazole and so may represent H2B in transcriptionally active chromatin. The slowly exchanging fraction was probably associated with chromatin domains surrounding active units. H3-GFP and H4-GFP were assembled into chromatin when DNA was replicated, and then >80% remained bound permanently. These results reveal that the inner core of the nucleosome is very stable, whereas H2B on the surface of active nucleosomes exchanges continually.

MeSH Terms
Cell Line Cell Nucleus/metabolism,ultrastructure Chromatin/metabolism,ultrastructure Chromosomes/metabolism,ultrastructure DNA/metabolism Dichlororibofuranosylbenzimidazole/pharmacology Green Fluorescent Proteins HeLa Cells Histones/genetics,metabolism Humans Kinetics Luminescent Proteins/genetics Macromolecular Substances Microscopy, Fluorescence/methods Nucleosomes/metabolism Photochemistry Protein Binding/drug effects,physiology Recombinant Fusion Proteins/genetics,metabolism S Phase/physiology Transfection
Chemicals
Chromatin Histones Luminescent Proteins Macromolecular Substances Nucleosomes Recombinant Fusion Proteins Green Fluorescent Proteins Dichlororibofuranosylbenzimidazole DNA
Authors & Affiliations
2 authors, click to expand affiliations / ORCID
Kimura H
Sir William Dunn School of Pathology, University of Oxford, Oxford OX1 3RE, United Kingdom.
Cook P R
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Article Info
Journal
The Journal of cell biology
Abbr.
J Cell Biol
ISSN
0021-9525
DOI
10.1083/jcb.153.7.1341
Published
2001-06-25
Pages
1341-53
Language
English
Region
United States
NLM ID
0375356
PMCID
PMC2150718
Subset
IM
External Links
PubMed source DOI: 10.1083/jcb.153.7.1341 PMC Full Text View journal details
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