In Vitro Assembly of the CENP-B/alpha-satellite DNA/core Histone Complex: CENP-B Causes Nucleosome Positioning

Genes Cells. 1998 Aug;3(8):533-48. doi: 10.1046/j.1365-2443.1998.00210.x.

Abstract

Background: We have studied the nucleosome structure formed from alpha-satellite DNA bound with CENP-B and core histones, in order to develop a previous proposal that the CENP-B dimer may play a critical role in the assembly of higher order structures of the human centromere by juxtaposing CENP-B boxes in long alpha-satellite arrays.

Results: The dimeric structure of CENP-B was sufficiently stable to bundle together two 3.5 kbp DNA fragments when each DNA contained a CENP-B box. When the same length of DNA included two CENP-B boxes, the intra-molecular interaction with the CENP-B dimer predominated, resulting in the formation of loop structures. The in vitro assembly of CENP-B/alpha-satellite DNA/core histone complexes with the aid of nucleosome assembly protein-1 (NAP-1) permitted an investigation into the nucleosome arrangement in alpha-satellite DNA with CENP-B bound to CENP-B boxes. Footprint analyses with micrococcal nuclease (MNase) revealed that CENP-B causes nucleosome positioning between pairs of CENP-B boxes with unique hypersensitive sites created on both sides.

Conclusion: We propose that CENP-B functions as a structural factor in the centromere region in order to establish a unique, centromere specific pattern of nucleosome positioning.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Autoantigens*
  • Base Sequence
  • Binding Sites
  • Cell Cycle Proteins
  • Centromere / metabolism*
  • Centromere / ultrastructure
  • Centromere Protein B
  • Chromatin / metabolism
  • Chromatin / ultrastructure
  • Chromosomal Proteins, Non-Histone / metabolism*
  • DNA / ultrastructure
  • DNA Footprinting
  • DNA, Satellite / metabolism*
  • DNA-Binding Proteins*
  • Dimerization
  • Histones / metabolism*
  • Humans
  • Models, Molecular
  • Molecular Sequence Data
  • Nuclear Proteins
  • Nucleic Acid Conformation
  • Nucleosome Assembly Protein 1
  • Nucleosomes / metabolism*
  • Protein Binding
  • Proteins / metabolism

Substances

  • Autoantigens
  • CENPB protein, human
  • Cell Cycle Proteins
  • Centromere Protein B
  • Chromatin
  • Chromosomal Proteins, Non-Histone
  • DNA, Satellite
  • DNA-Binding Proteins
  • Histones
  • NAP1L1 protein, human
  • Nuclear Proteins
  • Nucleosome Assembly Protein 1
  • Nucleosomes
  • Proteins
  • DNA