Cell-cycle-dependent structural transitions in the human CENP-A nucleosome in vivo.

Laboratory of Receptor Biology and Gene Expression, National Cancer Institute, NIH, Bethesda, MD 20892, USA.
Cell (Impact Factor: 31.96). 07/2012; 150(2):317-26. DOI: 10.1016/j.cell.2012.05.035
Source: PubMed

ABSTRACT In eukaryotes, DNA is packaged into chromatin by canonical histone proteins. The specialized histone H3 variant CENP-A provides an epigenetic and structural basis for chromosome segregation by replacing H3 at centromeres. Unlike exclusively octameric canonical H3 nucleosomes, CENP-A nucleosomes have been shown to exist as octamers, hexamers, and tetramers. An intriguing possibility reconciling these observations is that CENP-A nucleosomes cycle between octamers and tetramers in vivo. We tested this hypothesis by tracking CENP-A nucleosomal components, structure, chromatin folding, and covalent modifications across the human cell cycle. We report that CENP-A nucleosomes alter from tetramers to octamers before replication and revert to tetramers after replication. These structural transitions are accompanied by reversible chaperone binding, chromatin fiber folding changes, and previously undescribed modifications within the histone fold domains of CENP-A and H4. Our results reveal a cyclical nature to CENP-A nucleosome structure and have implications for the maintenance of epigenetic memory after centromere replication.

  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: The assembly of centromeric nucleosomes is mediated by histone variant-specific chaperones. In budding yeast, the centromere-specific histone H3 variant is Cse4, and the histone chaperone Scm3 functions as a Cse4-specific nucleosome assembly factor. Here, we show that Scm3 exhibits specificity for Cse4-H4, but also interacts with major-type H3-H4 and H2A-H2B. Previously published structures of the Scm3 histone complex demonstrate that Scm3 binds only one copy of Cse4-H4. Consistent with this, we show that Scm3 deposits Cse4-H4 through a dimer intermediate onto deoxyribonucleic acid (DNA) to form a (Cse4-H4)2-DNA complex (tetrasome). Scm3-bound Cse4-H4 does not form a tetramer in the absence of DNA. Moreover, we demonstrate that Cse4 and H3 are structurally compatible to be incorporated in the same nucleosome to form heterotypic particles. Our data shed light on the mechanism of Scm3-mediated nucleosome assembly at the centromere.
    Nucleic Acids Research 03/2014; · 8.81 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: To maintain genome function and stability, DNA sequence and its organization into chromatin must be duplicated during cell division. Understanding how entire chromosomes are copied remains a major challenge. Here, we use nascent chromatin capture (NCC) to profile chromatin proteome dynamics during replication in human cells. NCC relies on biotin-dUTP labelling of replicating DNA, affinity purification and quantitative proteomics. Comparing nascent chromatin with mature post-replicative chromatin, we provide association dynamics for 3,995 proteins. The replication machinery and 485 chromatin factors such as CAF-1, DNMT1 and SUV39h1 are enriched in nascent chromatin, whereas 170 factors including histone H1, DNMT3, MBD1-3 and PRC1 show delayed association. This correlates with H4K5K12diAc removal and H3K9me1 accumulation, whereas H3K27me3 and H3K9me3 remain unchanged. Finally, we combine NCC enrichment with experimentally derived chromatin probabilities to predict a function in nascent chromatin for 93 uncharacterized proteins, and identify FAM111A as a replication factor required for PCNA loading. Together, this provides an extensive resource to understand genome and epigenome maintenance.
    Nature Cell Biology 02/2014; · 20.76 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Genetic information stored in DNA is accurately copied and transferred to subsequent generations through DNA replication. This process is accomplished through the concerted actions of highly conserved DNA replication components. Epigenetic information stored in the form of histone modifications and DNA methylation, constitutes a second layer of regulatory information important for many cellular processes, such as gene expression regulation, chromatin organization, and genome stability. During DNA replication, epigenetic information must also be faithfully transmitted to subsequent generations. How this monumental task is achieved remains poorly understood. In this review, we will discuss recent advances on the role of DNA replication components in the inheritance of epigenetic marks, with a particular focus on epigenetic regulation in fission yeast. Based on these findings, we propose that specific DNA replication components function as key regulators in the replication of epigenetic information across the genome.
    Protein & Cell 04/2014; · 3.22 Impact Factor


Available from
May 19, 2014