CENP-A nucleosome—a chromatin-embedded pedestal for the centromere: lessons learned from structural biology

Ali-Ahmad, Ahmad; Sekulić, Nikolina

doi:10.1042/ebc20190074

Cited by 17 publications

(7 citation statements)

References 137 publications

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“…CENP-A, when present, forms a nucleosome structure in which the canonical histone H3 is replaced with CENP-A. Much debate has surrounded as to what makes the CENP-A chromatin “special,” summarized in other studies ( McKinley and Cheeseman, 2016 ; Ali-Ahmad and Sekulić, 2020 ; Mitra et al, 2020a ). CCAN components CENP-C and CENP-N in vertebrates directly bind to the CENP-A nucleosome to assemble the whole CCAN structure in the centromeric chromatin ( Carroll et al, 2009 ; Kato et al, 2013 ; Watanabe et al, 2019 ; Ariyoshi et al, 2021 ), and CENP-U Ame1 -Q Okp1 in budding yeast also directly binds to the CENP-A Cse4 nucleosome ( Anedchenko et al, 2019 ; Fischböck-Halwachs et al, 2019 ).…”

Section: Inner Kinetochore and The Ccanmentioning

confidence: 99%

Kinetochore Architecture Employs Diverse Linker Strategies Across Evolution

Sridhar

Fukagawa

2022

Front. Cell Dev. Biol.

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The assembly of a functional kinetochore on centromeric chromatin is necessary to connect chromosomes to the mitotic spindle, ensuring accurate chromosome segregation. This connecting function of the kinetochore presents multiple internal and external structural challenges. A microtubule interacting outer kinetochore and centromeric chromatin interacting inner kinetochore effectively confront forces from the external spindle and centromere, respectively. While internally, special inner kinetochore proteins, defined as “linkers,” simultaneously interact with centromeric chromatin and the outer kinetochore to enable association with the mitotic spindle. With the ability to simultaneously interact with outer kinetochore components and centromeric chromatin, linker proteins such as centromere protein (CENP)-C or CENP-T in vertebrates and, additionally CENP-QOkp1-UAme1 in yeasts, also perform the function of force propagation within the kinetochore. Recent efforts have revealed an array of linker pathways strategies to effectively recruit the largely conserved outer kinetochore. In this review, we examine these linkages used to propagate force and recruit the outer kinetochore across evolution. Further, we look at their known regulatory pathways and implications on kinetochore structural diversity and plasticity.

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Section: Inner Kinetochore and The Ccanmentioning

confidence: 99%

Kinetochore Architecture Employs Diverse Linker Strategies Across Evolution

Sridhar

Fukagawa

2022

Front. Cell Dev. Biol.

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“…Within these centromeric regions, CENP-A establishes the centromere that recruits the kinetochore 12 – 14 , a large protein complex required to bridge the chromosomes with the spindle microtubules. CENP-A exhibits a number of structural differences compared to canonical H3 15 . A loop region in CENP-A protrudes from the nucleosome, exposing residues arginine 80 and glycine 81 (RG-loop) and thereby creating a site for CENP-N recruitment 16 – 18 , albeit this role of the RG-loop has been challenged by recent cryoEM structures of the inner centromere 19 , 20 (Fig.…”

Section: Introductionmentioning

confidence: 99%

CENP-A and CENP-B collaborate to create an open centromeric chromatin state

Nagpal,

Ali-Ahmad,

Hirano

et al. 2023

Nat Commun

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Centromeres are epigenetically defined via the presence of the histone H3 variant CENP-A. Contacting CENP-A nucleosomes, the constitutive centromere associated network (CCAN) and the kinetochore assemble, connecting the centromere to spindle microtubules during cell division. The DNA-binding centromeric protein CENP-B is involved in maintaining centromere stability and, together with CENP-A, shapes the centromeric chromatin state. The nanoscale organization of centromeric chromatin is not well understood. Here, we use single-molecule fluorescence and cryoelectron microscopy (cryoEM) to show that CENP-A incorporation establishes a dynamic and open chromatin state. The increased dynamics of CENP-A chromatin create an opening for CENP-B DNA access. In turn, bound CENP-B further opens the chromatin fiber structure and induces nucleosomal DNA unwrapping. Finally, removal of CENP-A increases CENP-B mobility in cells. Together, our studies show that the two centromere-specific proteins collaborate to reshape chromatin structure, enabling the binding of centromeric factors and establishing a centromeric chromatin state.

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“…Centromeres are the chromosomal sites for assembly of kinetochores, the fundamental complex necessary for proper chromosome segregation in both meiosis and mitosis [ 1 , 2 ]. In humans they are composed of highly repetitive arrays of alpha satellite DNA (α-sat) that stretches over megabase-long regions [ 3 ].…”

Section: Introductionmentioning

confidence: 99%

“…Centromeric DNA and its DNA binding protein CENP-B have been recently implicated in centromere stability or function [ 1 , 8 – 12 ]. Yet, the repetitive nature of these loci has hindered their detailed molecular characterization.…”

Section: Introductionmentioning

confidence: 99%

Enrichment of centromeric DNA from human cells

et al. 2022

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Centromeres are key elements for chromosome segregation. Canonical centromeres are built over long-stretches of tandem repetitive arrays. Despite being quite abundant compared to other loci, centromere sequences overall still represent only 2 to 5% of the human genome, therefore studying their genetic and epigenetic features is a major challenge. Furthermore, sequencing of centromeric regions requires high coverage to fully analyze length and sequence variations, and this can be extremely costly. To bypass these issues, we have developed a technique, named CenRICH, to enrich for centromeric DNA from human cells based on selective restriction digestion and size fractionation. Combining restriction enzymes cutting at high frequency throughout the genome, except within most human centromeres, with size-selection of fragments >20 kb, resulted in over 25-fold enrichment in centromeric DNA. High-throughput sequencing revealed that up to 60% of the DNA in the enriched samples is made of centromeric repeats. We show that this method can be used in combination with long-read sequencing to investigate the DNA methylation status of certain centromeres and, with a specific enzyme combination, also of their surrounding regions (mainly HSATII). Finally, we show that CenRICH facilitates single-molecule analysis of replicating centromeric fibers by DNA combing. This approach has great potential for making sequencing of centromeric DNA more affordable and efficient and for single DNA molecule studies.

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CENP-A nucleosome—a chromatin-embedded pedestal for the centromere: lessons learned from structural biology

Cited by 17 publications

References 137 publications

Kinetochore Architecture Employs Diverse Linker Strategies Across Evolution

Kinetochore Architecture Employs Diverse Linker Strategies Across Evolution

CENP-A and CENP-B collaborate to create an open centromeric chromatin state

Enrichment of centromeric DNA from human cells

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