ReviewEpigenetic specification of centromeres by CENP-A
Introduction
The centromere/kinetochore complex orchestrates chromosome segregation and, along with telomeres and origins of replication, constitutes a key element to ensure faithful propagation of the genome. After the cloning of the first centromere sequence from Saccharomyces cerevisiae by Clarke and Carbon [1], many researchers tried to identify a similar “magic sequence” in other organisms. However, the complexity of centromere sequences became clear upon their discovery in Schizosaccharomyces pombe [2], [3]. Fission yeast centromeres are 50–100 kb long, in contrast to 125 bp in budding yeast, and contain a complex arrangement of repeated sequences. This difference in sequence organization led Pluta et al. to distinguish between point centromeres (those of S. cerevisiae) and regional centromeres (those of S. pombe) [4]. The presence of repeated sequences appears to be a characteristic of most centromeres analyzed to date from plants, flies and humans [5], [6], [7]. Importantly, centromeres form at a subset of these repetitive DNA arrays and thus the site of kinetochore assembly appears to be determined by factors other than DNA sequence alone. Indeed, early studies revealed the epigenetic nature of centromeres. In dicentric chromosomes (i.e., carrying two centromeres) only one of them is active for recruitment of kinetochore proteins while the other is inactivated [8], [9]. Steiner and Clarke [10] showed that a nonfunctional centromere present on a circular minichromosome could be converted to a functional one without changes in the content, structural arrangement or chemical modification state of the DNA. Strong evidence supporting that a particular DNA sequence is neither necessary nor sufficient for centromere specification comes from the existence of neocentromeres that arise at otherwise acentric fragments resulting from a chromosome rearrangement. These neocentromeres are devoid of the repeated sequences found at canonical centromeres but recruit most of the centromeric and pericentromeric proteins required for proper chromosome segregation [11].
What, if not DNA sequence, determines the site of kinetochore assembly? Increasing experimental evidence suggests that chromatin composition and organization play a major role in centromere specification and propagation [12], [13], [14]. Centromere Protein A, CENP-A, is a histone H3 variant that replaces canonical histone H3 in the centromeric nucleosomes of all eukaryotes [15], [16], [17], [18], [19]. Inactivation or down regulation of CENP-A in different experimental systems results in chromosome segregation defects and eventually cell death, and its presence is required for assembly of all other centromeric proteins [20], [21], [22], [23], [24], [25], [26]. It has been therefore proposed that CENP-A is the epigenetic mark of the centromere. Although we are far from understanding the molecular determinants of this mark and its inheritance, recent studies have provided exciting clues that we summarize in this review.
Section snippets
What makes CENP-A unique?
CENP-A localizes specifically at active centromeres, although its overexpression can lead to promiscuous localization all over the chromosomes [27], [28], [29]. This 17-kDa histone variant displays over 60% sequence identity to canonical H3 over the C-terminal domain that includes the “histone fold” domain (HFD) [15]. In contrast, the N-terminal region of CENP-A shares the flexible nature of histone tails but shows little amino acid sequence similarity to H3 or any other histone variant, and it
When does CENP-A deposition occur?
Maintenance of centromere identity requires incorporation of new CENP-A during or after replication of centromeric DNA. In S. cerevisiae, all pre-existing CENP-A is replaced by newly synthesized CENP-A during S phase [58] whereas in S. pombe, two pathways of CENP-A deposition exist at different times of the cell cycle, S phase and G2 [59], [60]. Experiments in Arabidopsis thaliana suggest that most CENP-A is loaded in G2 by a replication-independent mechanism [61]. Early studies in human cells
Histone chaperones and CENP-A deposition
Despite their sequence and structural similarities, histone H3 variants are deposited by distinct chaperones: histone H3.1 is deposited by the Chromatin Assembly Factor 1 (CAF-1) complex during replication and DNA damage processing, while histone H3.3 is deposited at active chromatin by a complex containing the HIRA protein in a process independent of DNA synthesis [78]. Some evidences suggested that these two complexes may contribute to CENP-A deposition in yeast, but neither of them could be
Conclusions/perspectives
Our knowledge on how centromere identity is propagated from one generation to the next has increased enormously over the last decade. Many factors have been identified that interact with CENP-A physically and/or genetically, and a number of them have been shown to affect the incorporation or stabilization of CENP-A at centromeres (Fig. 1). However, the exact role of most of these factors remains unknown. The development of an in vitro assay for CENP-A incorporation from purified components
Acknowledgments
We thank Juan Méndez (CNIO) and members of the lab for critically reading the manuscript, and colleagues in the field for helpful discussions. We apologize to those whose work is not cited due to space limitations. Research in our lab is supported by the Spanish Ministry of Science and Innovation (grants BFU2007-66627 and CSD2007-0015, and FPI predoctoral fellowship to P.S.), Fundación Caja Madrid, the Epigenome Network of Excellence (EU) and EMBO (postdoctoral fellowship ALTF 77-2007 to R.B.).
References (109)
- et al.
Composite motifs and repeat symmetry in S. pombe centromeres: direct analysis by integration of NotI restriction sites
Cell
(1989) - et al.
Molecular structure of a functional Drosophila centromere
Cell
(1997) - et al.
A novel epigenetic effect can alter centromere function in fission yeast
Cell
(1994) - et al.
Neocentromeres: new insights into centromere structure, disease development, and karyotype evolution
Am. J. Hum. Genet.
(2008) - et al.
Immunolocalization of CENP-A suggests a distinct nucleosome structure at the inner kinetochore plate of active centromeres
Curr. Biol.
(1997) - et al.
Inactivation of a human kinetochore by specific targeting of chromatin modifiers
Dev. Cell.
(2008) - et al.
Cse4p is a component of the core centromere of Saccharomyces cerevisiae
Cell
(1998) - et al.
Mislocalization of the Drosophila centromere-specific histone CID promotes formation of functional ectopic kinetochores
Dev. Cell.
(2006) - et al.
Centromeric chromatin: what makes it unique?
Curr. Opin. Genet. Dev.
(2005) - et al.
Centromere identity maintained by nucleosomes assembled with histone H3 containing the CENP-A targeting domain
Mol. Cell.
(2007)
Centromeric nucleosomes induce positive DNA supercoils
Cell
CENP-A-containing nucleosomes: easier disassembly versus exclusive centromeric localization
J. Mol. Biol.
Nonhistone Scm3 and histones CenH3-H4 assemble the core of centromere-specific nucleosomes
Cell
Scm3 is essential to recruit the histone h3 variant cse4 to centromeres and to maintain a functional kinetochore
Mol. Cell.
Fission yeast Scm3: a CENP-A receptor required for integrity of subkinetochore chromatin
Mol. Cell.
Fission yeast Scm3 mediates stable assembly of Cnp1/CENP-A into centromeric chromatin
Mol. Cell.
Conserved organization of centromeric chromatin in flies and humans
Dev. Cell.
A new fluorescence resonance energy transfer approach demonstrates that the histone variant H2AZ stabilizes the histone octamer within the nucleosome
J. Biol. Chem.
Stable kinetochore-microtubule attachment constrains centromere positioning in metaphase
Curr. Biol.
(N1/N2)-related protein, Sim3, binds CENP-A and is required for its deposition at fission yeast centromeres
Mol. Cell.
Incorporation of Drosophila CID/CENP-A and CENP-C into centromeres during early embryonic anaphase
Curr. Biol.
CCAN makes multiple contacts with centromeric DNA to provide distinct pathways to the outer kinetochore
Cell
Proteolysis contributes to the exclusive centromere localization of the yeast Cse4/CENP-A histone H3 variant
Curr. Biol.
HJURP is a cell-cycle-dependent maintenance and deposition factor of CENP-A at centromeres
Cell
Centromere-specific assembly of CENP-a nucleosomes is mediated by HJURP
Cell
Histone H3.1 and H3.3 complexes mediate nucleosome assembly pathways dependent or independent of DNA synthesis
Cell
Mis16 and Mis18 are required for CENP-A loading and histone deacetylation at centromeres
Cell
Mis6, a fission yeast inner centromere protein, acts during G1/S and forms specialized chromatin required for equal segregation
Cell
Xenopus CENP-A assembly into chromatin requires base excision repair proteins
DNA. Repair. (Amst).
Common ancestry of the CENP-A chaperones Scm3 and HJURP
Cell
Priming of centromere for CENP-A recruitment by human hMis18alpha, hMis18beta, and M18BP1
Dev. Cell.
CENP-B controls centromere formation depending on the chromatin context
Cell
Isolation of a yeast centromere and construction of functional small circular chromosomes
Nature
Analysis of centromeric DNA in the fission yeast Schizosaccharomyces pombe
Proc. Natl. Acad. Sci. U. S. A.
The centromere: hub of chromosomal activities
Science
Genomic and genetic definition of a functional human centromere
Science
Genetic definition and sequence analysis of Arabidopsis centromeres
Science
Three related centromere proteins are absent from the inactive centromere of a stable isodicentric chromosome
Chromosoma
A dicentric chromosome of Drosophila melanogaster showing alternate centromere inactivation
Chromosoma
Formation of functional centromeric chromatin is specified epigenetically in Candida albicans
Proc. Natl. Acad. Sci. U. S. A.
A 17-kD centromere protein (CENP-A) copurifies with nucleosome core particles and with histones
J. Cell. Biol.
Human CENP-A contains a histone H3 related histone fold domain that is required for targeting to the centromere
J. Cell. Biol.
A histone-H3-like protein in C. elegans
Nature
Heterochromatic deposition of centromeric histone H3-like proteins
Proc. Natl. Acad. Sci. U. S. A.
A mutation in CSE4, an essential gene encoding a novel chromatin-associated protein in yeast, causes chromosome nondisjunction and cell cycle arrest at mitosis
Genes Dev.
Requirement of Mis6 centromere connector for localizing a CENP-A-like protein in fission yeast
Science
Early disruption of centromeric chromatin organization in centromere protein A (Cenpa) null mice
Proc. Natl. Acad. Sci. U. S. A.
Functional analysis of kinetochore assembly in Caenorhabditis elegans
J. Cell. Biol.
The role of Drosophila CID in kinetochore formation, cell-cycle progression and heterochromatin interactions
Nat. Cell. Biol.
CENP-A is required for accurate chromosome segregation and sustained kinetochore association of BubR1
Mol. Cell. Biol.
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2013, Molecular and Biochemical ParasitologyCitation Excerpt :CENP-A (CENH3), the centromeric histone is an inner kinetochore component which maintains the centromere identity and function throughout the cell cycle by replacing core histone H3 containing nucleosomes at the centromeric chromatin [1–10].
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2010, Current Opinion in Cell BiologyCitation Excerpt :When introduced into fly cells, yeast Cse4, human CENP-A, and worm HCP-3 localize to Drosophila centromeres [7]; Cse4 can localize to and functionally complement human CENP-A depletion [8]; residues of CENH3 swapped into H3 allow the chimera to localize to centromeres [9,10] and, avian CENP-A can preferentially associate with mouse 120 bp centromeric repeats [11]. Consequently, studies dissecting CENH3 assembly, localization, and structure are of considerable interest to achieve a unitary model for centromere function [12,13]. Chaperones guiding CENH3 assembly to centromeres have been pursued for over a decade.
Building centromeres: Home sweet home or a nomadic existence?
2010, Current Opinion in Genetics and DevelopmentCitation Excerpt :What does CENP-A chromatin look like once it is established and operating in the context of a functional centromere? The structure and assembly of CENP-A chromatin are highly active areas of research and several excellent reviews have been recently published [5,41,42], and we will therefore only summarise the state of play. Evidence has been presented for, and in some cases, against, a variety of CENP-A nucleosome structures, including regular homotypic octamers, hexamers, hemisomes, and right-handed wrapping of DNA around CENP-A nucleosomes [41,43–48].