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Nucleic Acids Res. 2014 Oct;42(18):11502-16. doi: 10.1093/nar/gku835. Epub 2014 Sep 16.

Replication of alpha-satellite DNA arrays in endogenous human centromeric regions and in human artificial chromosome.

Author information

1
Developmental Therapeutics Branch, National Cancer Institute, National Institute of Health, Bethesda, MD, 20892, USA.
2
Laboratory of Cell Engineering, Department of Frontier Research, Kazusa DNA Research Institute, Kisarazu, Chiba, 292-0818, Japan.
3
Center for Biomolecular Science and Engineering, University of California, Santa Cruz, CA, 95064, USA.
4
Wellcome Trust Centre for Cell Biology, University of Edinburgh, Edinburgh, EH9 3JR, UK.
5
Developmental Therapeutics Branch, National Cancer Institute, National Institute of Health, Bethesda, MD, 20892, USA kouprinn@mail.nih.gov.

Abstract

In human chromosomes, centromeric regions comprise megabase-size arrays of 171 bp alpha-satellite DNA monomers. The large distances spanned by these arrays preclude their replication from external sites and imply that the repetitive monomers contain replication origins. However, replication within these arrays has not previously been profiled and the role of alpha-satellite DNA in initiation of DNA replication has not yet been demonstrated. Here, replication of alpha-satellite DNA in endogenous human centromeric regions and in de novo formed Human Artificial Chromosome (HAC) was analyzed. We showed that alpha-satellite monomers could function as origins of DNA replication and that replication of alphoid arrays organized into centrochromatin occurred earlier than those organized into heterochromatin. The distribution of inter-origin distances within centromeric alphoid arrays was comparable to the distribution of inter-origin distances on randomly selected non-centromeric chromosomal regions. Depletion of CENP-B, a kinetochore protein that binds directly to a 17 bp CENP-B box motif common to alpha-satellite DNA, resulted in enrichment of alpha-satellite sequences for proteins of the ORC complex, suggesting that CENP-B may have a role in regulating the replication of centromeric regions. Mapping of replication initiation sites in the HAC revealed that replication preferentially initiated in transcriptionally active regions.

PMID:
25228468
PMCID:
PMC4191410
DOI:
10.1093/nar/gku835
[Indexed for MEDLINE]
Free PMC Article

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