Effects of H1 Histone Variant Overexpression on Chromatin Structure*
- From the Departments of ‡Biochemistry and§Physiology, University of Mississippi Medical Center, Jackson, Mississippi 39216-4505
Abstract
The importance of histone H1 heterogeneity and total H1 stoichiometry in chromatin has been enigmatic. Here we report a detailed characterization of the chromatin structure of cells overexpressing either H10 or H1c. Nucleosome spacing was found to change during cell cycle progression, and overexpression of either variant in exponentially growing cells results in a 15-base pair increase in nucleosome repeat length. H1 histones can also assemble on chromatin and influence nucleosome spacing in the absence of DNA replication. Overexpression of H10 and, to a lesser extent, H1c results in a decreased rate of digestion of chromatin by micrococcal nuclease. Using green fluorescent protein-tagged H1 variants, we show that micrococcal nuclease-resistant chromatin is specifically enriched in the H10 variant. Overexpression of H10 results in the appearance of a unique mononucleosome species of higher mobility on nucleoprotein gels. Domain switch mutagenesis revealed that either the N-terminal tail or the central globular domain of the H10 protein could independently give rise to this unique mononucleosome species. These results in part explain the differential effects of H10 and H1c in regulating chromatin structure and function.
Footnotes
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↵* This work was supported by Grant MCB-9305308 from the National Science Foundation, an institutional grant from the University of Mississippi Medical Center, and a donation from the F. D. Wade Research Fund.The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
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↵¶ To whom correspondence should be addressed: Dept. of Biochemistry, University of Mississippi Medical Center, 2500 N. State St., Jackson, MS 39216-4505. Tel.: 601-984-1848; Fax: 601-984-1501; E-mail: dsittman@biochem.umsmed.edu.
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↵2 A. Gunjan, B. T. Alexander, D. B. Sittman, and D. T. Brown, unpublished observations.
- Abbreviations:
- bp
-
base pair(s)
- MN
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micrococcal nuclease
- GFP
-
green fluorescent protein
- HPLC
-
high performance liquid chromatography
- NP
-
nucleoprotein
- EtBr
-
ethidium bromide
- HU
-
hydroxyurea
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- Received April 26, 1999.
- Revision received September 29, 1999.
- The American Society for Biochemistry and Molecular Biology, Inc.
