Abstract
BackgroundChromatin containing the histone variant CENP-A (CEN chromatin) exists as an essential domain at every centromere and heritably marks the location of kinetochore assembly. The size of the CEN chromatin domain on alpha satellite DNA in humans has been shown to vary according to underlying array size. However, the average amount of CENP-A reported at human centromeres is largely consistent, implying the genomic extent of CENP-A chromatin domains more likely reflects variations in the number of CENP-A subdomains and/or the density of CENP-A nucleosomes within individual subdomains. Defining the organizational and spatial properties of CEN chromatin would provide insight into centromere inheritance via CENP-A loading in G1 and the dynamics of its distribution between mother and daughter strands during replication.ResultsUsing a multi-color protein strategy to detect distinct pools of CENP-A over several cell cycles, we show that nascent CENP-A is equally distributed to sister centromeres. CENP-A distribution is independent of previous or subsequent cell cycles in that centromeres showing disproportionately distributed CENP-A in one cycle can equally divide CENP-A nucleosomes in the next cycle. Furthermore, we show using extended chromatin fibers that maintenance of the CENP-A chromatin domain is achieved by a cycle-specific oscillating pattern of new CENP-A nucleosomes next to existing CENP-A nucleosomes over multiple cell cycles. Finally, we demonstrate that the size of the CENP-A domain does not change throughout the cell cycle and is spatially fixed to a similar location within a given alpha satellite DNA array.ConclusionsWe demonstrate that most human chromosomes share similar patterns of CENP-A loading and distribution and that centromere inheritance is achieved through specific placement of new CENP-A near existing CENP-A as assembly occurs each cell cycle. The loading pattern fixes the location and size of the CENP-A domain on individual chromosomes. These results suggest that spatial and temporal dynamics of CENP-A are important for maintaining centromere identity and genome stability.Electronic supplementary materialThe online version of this article (doi:10.1186/s13072-016-0071-7) contains supplementary material, which is available to authorized users.
Highlights
Chromatin containing the histone variant centromere protein A (CENP-A) (CEN chromatin) exists as an essential domain at every centromere and heritably marks the location of kinetochore assembly
Similar CENP‐A nucleosome quantities are maintained on several chromosomes Prior studies have suggested that amounts of CENPA vary among human centromeres and that CENP-A nucleosomes segregate randomly between daughter chromatids during replication [23, 24]
The assembly of CENP-A nucleosomes into the CEN chromatin domain that is faithfully maintained is important for genome and chromosome stability
Summary
Chromatin containing the histone variant CENP-A (CEN chromatin) exists as an essential domain at every centromere and heritably marks the location of kinetochore assembly. The size of the CEN chromatin domain on alpha satellite DNA in humans has been shown to vary according to underlying array size. Alpha satellite forms a homogenous array extending over megabases on a given chromosome, centromere proteins normally occupy only a portion of the array [10,11,12,13]. This CENP-A/centromeric (CEN) chromatin domain is uniquely arranged as interspersed subdomains of CENP-A and H3 nucleosomes, including both H3.1 and H3.3 [13,14,15,16,17]. A range of CENP-A domain sizes among human centromeres suggests that the organization of CEN chromatin, such as the concentration of CENP-A nucleosomes within subdomains, varies based on the total genomic extent of the CEN chromatin domain
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