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Updated: Mar 16, 2026

Immunofluorescence Analysis of Endogenous and Exogenous Centromere-kinetochore Proteins
Published on: March 3, 2016
The Flexible Ends of CENP-A Nucleosome Are Required for Mitotic Fidelity
Yohan Roulland1, Khalid Ouararhni2, Mladen Naidenov3
1Université de Grenoble Alpes/INSERM U1209/CNRS UMR 5309, 38042 Grenoble Cedex 9, France.
Abstract:
CENP-A is a histone variant, which replaces histone H3 at centromeres and confers unique properties to centromeric chromatin. The crystal structure of CENP-A nucleosome suggests flexible nucleosomal DNA ends, but their dynamics in solution remains elusive and their implication in centromere function is unknown. Using electron cryo-microscopy, we determined the dynamic solution properties of the CENP-A nucleosome. Our biochemical, proteomic, and genetic data reveal that higher flexibility of DNA ends impairs histone H1 binding to the CENP-A nucleosome. Substituting the 2-turn αN-helix of CENP-A with the 3-turn αN-helix of H3 results in compact particles with rigidified DNA ends, able to bind histone H1. In vivo replacement of CENP-A with H3-CENP-A hybrid nucleosomes leads to H1 recruitment, delocalization of kinetochore proteins, and significant mitotic and cytokinesis defects. Our data reveal that the evolutionarily conserved flexible ends of the CENP-A nucleosomes are essential to ensure the fidelity of the mitotic pathway.
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