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Dev Cell. 2008 Apr;14(4):507-22. doi: 10.1016/j.devcel.2008.02.001.

Inactivation of a human kinetochore by specific targeting of chromatin modifiers.

Author information

1
Laboratory of Molecular Pharmacology, National Cancer Institute, National Institutes of Health, Building 37, Room 5040, 9000 Rockville Pike, Bethesda, MD 20892, USA.

Abstract

We have used a human artificial chromosome (HAC) to manipulate the epigenetic state of chromatin within an active kinetochore. The HAC has a dimeric alpha-satellite repeat containing one natural monomer with a CENP-B binding site, and one completely artificial synthetic monomer with the CENP-B box replaced by a tetracycline operator (tetO). This HAC exhibits normal kinetochore protein composition and mitotic stability. Targeting of several tet-repressor (tetR) fusions into the centromere had no effect on kinetochore function. However, altering the chromatin state to a more open configuration with the tTA transcriptional activator or to a more closed state with the tTS transcription silencer caused missegregation and loss of the HAC. tTS binding caused the loss of CENP-A, CENP-B, CENP-C, and H3K4me2 from the centromere accompanied by an accumulation of histone H3K9me3. Our results reveal that a dynamic balance between centromeric chromatin and heterochromatin is essential for vertebrate kinetochore activity.

PMID:
18410728
PMCID:
PMC2311382
DOI:
10.1016/j.devcel.2008.02.001
[Indexed for MEDLINE]
Free PMC Article

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