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Vol. 16, Issue 6, 2872-2881, June 2005

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Differential Subnuclear Localization and Replication Timing of Histone H3 Lysine 9 Methylation States

Rong Wu ^*, Anna V. Terry ^*, Prim B. Singh , and David M. Gilbert ^*

^* Department of Biochemistry and Molecular Biology, State University of New York Upstate Medical University, Syracuse, NY 13210; Research Center Borstel, D-23845 Borstel, Germany

Submitted November 16, 2004; Revised March 7, 2005; Accepted March 15, 2005
Monitoring Editor: Joseph Gall

Mono-, di-, and trimethylation of specific histone residues adds an additional level of complexity to the range of histone modifications that may contribute to a histone code. However, it has not been clear whether different methylated states reside stably at different chromatin sites or whether they represent dynamic intermediates at the same chromatin sites. Here, we have used recently developed antibodies that are highly specific for mono-, di-, and trimethylated lysine 9 of histone H3 (MeK9H3) to examine the subnuclear localization and replication timing of chromatin containing these epigenetic marks in mammalian cells. Me1K9H3 was largely restricted to early replicating, small punctate domains in the nuclear interior. Me2K9H3 was the predominant MeK9 epitope at the nuclear and nucleolar periphery and colocalized with sites of DNA synthesis primarily in mid-S phase. Me3K9H3 decorated late-replicating pericentric heterochromatin in mouse cells and sites of DAPI-dense intranuclear heterochromatin in human and hamster cells that replicated throughout S phase. Disruption of the Suv39h1,2 or G9a methyltransferases in murine embryonic stem cells resulted in a redistribution of methyl epitopes, but did not alter the overall spatiotemporal replication program. These results demonstrate that mono-, di-, and trimethylated states of K9H3 largely occupy distinct chromosome domains.

Address correspondence to: David M. Gilbert (gilbertd{at}upstate.edu).

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