Assembly in G1 phase and long-term stability are unique intrinsic features of CENP-A nucleosomes

Mol Biol Cell. 2013 Apr;24(7):923-32. doi: 10.1091/mbc.E13-01-0034. Epub 2013 Jan 30.

Abstract

Centromeres are the site of kinetochore formation during mitosis. Centromere protein A (CENP-A), the centromere-specific histone H3 variant, is essential for the epigenetic maintenance of centromere position. Previously we showed that newly synthesized CENP-A is targeted to centromeres exclusively during early G1 phase and is subsequently maintained across mitotic divisions. Using SNAP-based fluorescent pulse labeling, we now demonstrate that cell cycle-restricted chromatin assembly at centromeres is unique to CENP-A nucleosomes and does not involve assembly of other H3 variants. Strikingly, stable retention is restricted to the CENP-A/H4 core of the nucleosome, which we find to outlast general chromatin across several cell divisions. We further show that cell cycle timing of CENP-A assembly is independent of centromeric DNA sequences and instead is mediated by the CENP-A targeting domain. Unexpectedly, this domain also induces stable transmission of centromeric nucleosomes, independent of the CENP-A deposition factor HJURP. This demonstrates that intrinsic properties of the CENP-A protein direct its cell cycle-restricted assembly and induces quantitative mitotic transmission of the CENP-A/H4 nucleosome core, ensuring long-term stability and epigenetic maintenance of centromere position.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Autoantigens / genetics
  • Autoantigens / metabolism*
  • Cell Line
  • Centromere / genetics
  • Centromere / metabolism
  • Centromere Protein A
  • Chromosomal Proteins, Non-Histone / genetics
  • Chromosomal Proteins, Non-Histone / metabolism*
  • DNA Modification Methylases / genetics
  • DNA Modification Methylases / metabolism
  • DNA Repair Enzymes / genetics
  • DNA Repair Enzymes / metabolism
  • DNA-Binding Proteins / genetics
  • DNA-Binding Proteins / metabolism
  • G1 Phase*
  • HeLa Cells
  • Histones / genetics
  • Histones / metabolism
  • Humans
  • In Situ Hybridization, Fluorescence
  • Luminescent Proteins / genetics
  • Luminescent Proteins / metabolism
  • Microscopy, Fluorescence
  • Models, Genetic
  • Nucleosomes / genetics
  • Nucleosomes / metabolism*
  • Protein Stability
  • RNA Interference
  • Recombinant Fusion Proteins / genetics
  • Recombinant Fusion Proteins / metabolism
  • Time Factors
  • Tumor Suppressor Proteins / genetics
  • Tumor Suppressor Proteins / metabolism

Substances

  • Autoantigens
  • CENPA protein, human
  • Centromere Protein A
  • Chromosomal Proteins, Non-Histone
  • DNA-Binding Proteins
  • HJURP protein, human
  • Histones
  • Luminescent Proteins
  • Nucleosomes
  • Recombinant Fusion Proteins
  • Tumor Suppressor Proteins
  • DNA Modification Methylases
  • MGMT protein, human
  • DNA Repair Enzymes