Cell-Cycle-Coupled Structural Oscillation of Centromeric Nucleosomes in Yeast

The centromere is a specialized chromosomal structure that regulates chromosome segregation. Centromeres are marked by a histone H3 variant. In budding yeast, the histone H3 variant Cse4 is present in a single centromeric nucleosome. Experimental evidence supports several different models for the structure of centromeric nucleosomes. To investigate Cse4 copy number in live yeast, we developed a method coupling fluorescence correlation spectroscopy and calibrated imaging. We find that centromeric nucleosomes have one copy of Cse4 during most of the cell cycle, whereas two copies are detected at anaphase. The proposal of an anaphase-coupled structural change is supported by Cse4-Cse4 interactions, incorporation of Cse4, and the absence of Scm3 in anaphase. Nucleosome reconstitution and ChIP suggests both Cse4 structures contain H2A/H2B. The increase in Cse4 intensity and deposition at anaphase are also observed in Candida albicans. Our experimental evidence supports a cell-cycle-coupled oscillation of centromeric nucleosome structure in yeast. º A new method enables copy measurements of fluorescent proteins in live cells º Cse4-Cse4 interactions are restricted to anaphase at centromeres º Cse4 can be deposited at centromeres during anaphase º Scm3 is absent from Cse4 nucleosomes at anaphase In vivo single-molecule imaging of yeast cells reveals that centromeric nucleosomes oscillate between tetrameric and octameric forms through the cell cycle.