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Dev Cell. 2013 Mar 25;24(6):635-48. doi: 10.1016/j.devcel.2013.02.009. Epub 2013 Mar 14.

Chromosome engineering allows the efficient isolation of vertebrate neocentromeres.

Author information

1
Department of Molecular Genetics, National Institute of Genetics and The Graduate University for Advanced Studies SOKENDAI, Mishima, Shizuoka 411-8540, Japan.

Abstract

Centromeres are specified by sequence-independent epigenetic mechanisms in most organisms. Rarely, centromere repositioning results in neocentromere formation at ectopic sites. However, the mechanisms governing how and where neocentromeres form are unknown. Here, we established a chromosome-engineering system in chicken DT40 cells that allowed us to efficiently isolate neocentromere-containing chromosomes. Neocentromeres appear to be structurally and functionally equivalent to native centromeres. Chromatin immunoprecipitation sequencing (ChIP-seq) analysis with 18 neocentromeres revealed that the centromere-specific histone H3 variant CENP-A occupies an ∼40 kb region at each neocentromere, which has no preference for specific DNA sequence motifs. Furthermore, we found that neocentromeres were not associated with histone modifications H3K9me3, H3K4me2, and H3K36me3 or with early replication timing. Importantly, low but significant levels of CENP-A are detected around endogenous centromeres, which are capable of seeding neocentromere assembly if the centromere core is removed. In summary, our experimental system provides valuable insights for understanding how neocentromeres form.

PMID:
23499358
PMCID:
PMC3925796
DOI:
10.1016/j.devcel.2013.02.009
[Indexed for MEDLINE]
Free PMC Article

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