Sequence Dependent Nanoscale Structure of CENP-A Nucleosomes

CENP-A is a histone variant found in high abundance at the centromere. The centromere chromatin similar to the bulk chromatin consist of nucleosomes, but H3 histone of the bulk chromatin nucleosomes (H3 nucleosomes) is replaced with CENP-A histone. Additionally, the centromere comprises tandem repeats of α-satellite DNA which CENP-A nucleosomes assemble upon. However, the effect of the DNA sequence on the nucleosome assembly and centromere formation remains poorly understood. Here we investigated the structure of nucleosomes assembled with the CENP-A variant using Atomic Force Microscopy. We assembled both CENP-A nucleosomes and H3 nucleosomes on a DNA substrate containing an α-satellite motif and characterized their positioning and wrapping efficiency. We also studied CENP-A nucleosomes on the 601-positioning motif and non-specific DNA to compare their relative positioning and stability. CENP-A nucleosomes assembled on α-satellite DNA did not show any positional preference along the substrate, which is similar to H3 nucleosomes and CENP-A nucleosomes on non-specific DNA. The range of nucleosome wrapping efficiency was narrower on α-satellite DNA compared with non-specific DNA, suggesting a more stable complex. These findings indicate that DNA sequence and histone composition may be two of many factors required for accurate centromere assembly.