Abstract
CENP-A is an essential histone H3 variant that epigenetically marks the centromeric region of chromosomes. Here we show that CENP-A nucleosomes form characteristic clusters during the G1 phase of the cell cycle. 2D and 3D super-resolution microscopy and segmentation analysis reveal that these clusters encompass a globular rosette-like structure, which evolves into a more compact structure in late G1. The rosette-like clusters contain numerous CENP-A molecules and form a large cellular structure of ∼250–300 nm diameter with remarkably similar shapes for each centromere. Co-localization analysis shows that HJURP, the CENP-A chaperone, is located in the center of the rosette and serves as a nucleation point. The discovery of an HJURP-mediated CENP-A nucleation in human cells and its structural description provide important insights into the mechanism of CENP-A deposition and the organization of CENP-A chromatin in the centromeric region.
Highlights
CENP-A is an essential histone H3 variant that epigenetically marks the centromeric region of chromosomes
To analyze centromeric chromatin by super-resolution imaging, we have visualized the CENP-A nucleosomes by using an anti-CENP-A antibody coupled to Alexa Fluor conjugated secondary antibodies
We focused on the early G1 phase, because at this stage centromeric chromatin undergoes a major change due to the deposition of the newly synthesized CENP-A
Summary
CENP-A is an essential histone H3 variant that epigenetically marks the centromeric region of chromosomes. The discovery of an HJURP-mediated CENP-A nucleation in human cells and its structural description provide important insights into the mechanism of CENP-A deposition and the organization of CENP-A chromatin in the centromeric region. Human centromere DNA comprises 171 bp AT-rich alpha-satellite monomers that are arranged into tandem arrays[1,2]. An interesting aspect is that the “dilution” of CENP-A in daughter centromeres during S phase and its subsequent restoration at the G1 phase may be required for faithful cell division. In this context, a key question is how CENP-A is delivered to centromeric chromatin and maintained on centromeres during the cell cycle
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