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Chromosome Research

, 19:457 | Cite as

Genomic size of CENP-A domain is proportional to total alpha satellite array size at human centromeres and expands in cancer cells

  • Lori L. Sullivan
  • Christopher D. Boivin
  • Brankica Mravinac
  • Ihn Young Song
  • Beth A. Sullivan
Article

Abstract

Human centromeres contain multi-megabase-sized arrays of alpha satellite DNA, a family of satellite DNA repeats based on a tandemly arranged 171 bp monomer. The centromere-specific histone protein CENP-A is assembled on alpha satellite DNA within the primary constriction, but does not extend along its entire length. CENP-A domains have been estimated to extend over 2,500 kb of alpha satellite DNA. However, these estimates do not take into account inter-individual variation in alpha satellite array sizes on homologous chromosomes and among different chromosomes. We defined the genomic distance of CENP-A chromatin on human chromosomes X and Y from different individuals. CENP-A chromatin occupied different genomic intervals on different chromosomes, but despite inter-chromosomal and inter-individual array size variation, the ratio of CENP-A to total alpha satellite DNA size remained consistent. Changes in the ratio of alpha satellite array size to CENP-A domain size were observed when CENP-A was overexpressed and when primary cells were transformed by disrupting interactions between the tumor suppressor protein Rb and chromatin. Our data support a model for centromeric domain organization in which the genomic limits of CENP-A chromatin varies on different human chromosomes, and imply that alpha satellite array size may be a more prominent predictor of CENP-A incorporation than chromosome size. In addition, our results also suggest that cancer transformation and amounts of centromeric heterochromatin have notable effects on the amount of alpha satellite that is associated with CENP-A chromatin.

Keywords

Chromatin Chromosome Heterochromatin Histone Kinetochore Retinoblastoma 

Abbreviations

CENP-A

Centromere protein A

CCD

Charged coupled device

CHEF

Contour-clamped homogenous electric field

ChIP

Chromatin immunoprecipitation

CSK buffer

Cytoskeleton buffer

DNA

Deoxyribonucleic acid

E7

E7 protein subunit of HPV

FISH

Fluorescence in situ hybridization

FITC

Fluorescein isothiocyanate

FBS

Fetal bovine serum

HAT

Hypoxanthine/aminopterin/thymidine

HPV

Human papillomavirus

HOR

Higher order repeat

HSAX

Homo sapiens chromosome X

HSAY

Homo sapiens chromosome Y

Kb

Kilobase

MEM

Minimal essential medium

Mb

Megabase

PCR

Polymerase chain reaction

PVDF

Polyvinylidene fluoride

Rb

Retinoblastoma protein

RPMI

Roswell Park Memorial Institute medium

PFGE

Pulsed field gel electrophoresis

SDS

Sodium dodecyl sulfate

Notes

Acknowledgments

We thank Cyrus Vaziri (University of North Carolina, Chapel Hill) for providing cell lines DIP3 and DIP3-E7 and Chris Shaw for technical assistance. This work was supported in part by grants from the American Cancer Society (ACS IRG-72-001-29-IRG), March of Dimes Foundation (6-FY06-377 and 6-FY10-294), and NIH NIGMS (R01 GM069514) to BAS.

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Copyright information

© Springer Science+Business Media B.V. 2011

Authors and Affiliations

  • Lori L. Sullivan
    • 1
  • Christopher D. Boivin
    • 2
  • Brankica Mravinac
    • 1
    • 4
  • Ihn Young Song
    • 2
  • Beth A. Sullivan
    • 1
    • 3
  1. 1.Duke Institute for Genome Sciences & PolicyDuke UniversityDurhamUSA
  2. 2.Department of Genetics and GenomicsBoston University School of MedicineBostonUSA
  3. 3.Department of Molecular Genetics and MicrobiologyDuke University Medical CenterDurhamUSA
  4. 4.Division of Molecular BiologyRuder Boskovic InstituteZagrebCroatia

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