scholarly journals Diaphanous formin mDia2 regulates CENP-A levels at centromeres

2016 ◽  
Vol 213 (4) ◽  
pp. 415-424 ◽  
Author(s):  
Chenshu Liu ◽  
Yinghui Mao
Keyword(s):  
Quantitative Imaging ◽  
Active Form ◽  
Protein A ◽  
Small Gtpase ◽  
Genome Replication ◽  
Rac Gtpase ◽  
Epigenetic Marker ◽  
Centromere Protein A ◽  
Histone H3 Variant ◽  
Recognition Protein

Centromeres of higher eukaryotes are epigenetically defined by centromere protein A (CENP-A), a centromere-specific histone H3 variant. The incorporation of new CENP-A into centromeres to maintain the epigenetic marker after genome replication in S phase occurs in G1 phase; however, how new CENP-A is loaded and stabilized remains poorly understood. Here, we identify the formin mDia2 as essential for stable replenishment of new CENP-A at centromeres. Quantitative imaging, pulse-chase analysis, and high-resolution ratiometric live-cell studies demonstrate that mDia2 and its nuclear localization are required to maintain CENP-A levels at centromeres. Depletion of mDia2 results in a prolonged centromere association of holiday junction recognition protein (HJURP), the chaperone required for CENP-A loading. A constitutively active form of mDia2 rescues the defect in new CENP-A loading caused by depletion of male germ cell Rac GTPase-activating protein (MgcRacGAP), a component of the small GTPase pathway essential for CENP-A maintenance. Thus, the formin mDia2 functions downstream of the MgcRacGAP-dependent pathway in regulating assembly of new CENP-A containing nucleosomes at centromeres.

scholarly journals Guarding the Genome: CENP-A-Chromatin in Health and Cancer

Genes ◽  
2020 ◽  
Vol 11 (7) ◽  
pp. 810 ◽  
Author(s):  
Megan A. Mahlke ◽  
Yael Nechemia-Arbely
Keyword(s):  
Chromosome Segregation ◽  
Posttranslational Modifications ◽  
Current Knowledge ◽  
Protein A ◽  
Ctcf Binding ◽  
Centromere Protein A ◽  
Structure Specification ◽  
Binding Factor ◽  
Histone H3 Variant ◽  
Recognition Protein

Faithful chromosome segregation is essential for the maintenance of genomic integrity and requires functional centromeres. Centromeres are epigenetically defined by the histone H3 variant, centromere protein A (CENP-A). Here we highlight current knowledge regarding CENP-A-containing chromatin structure, specification of centromere identity, regulation of CENP-A deposition and possible contribution to cancer formation and/or progression. CENP-A overexpression is common among many cancers and predicts poor prognosis. Overexpression of CENP-A increases rates of CENP-A deposition ectopically at sites of high histone turnover, occluding CCCTC-binding factor (CTCF) binding. Ectopic CENP-A deposition leads to mitotic defects, centromere dysfunction and chromosomal instability (CIN), a hallmark of cancer. CENP-A overexpression is often accompanied by overexpression of its chaperone Holliday Junction Recognition Protein (HJURP), leading to epigenetic addiction in which increased levels of HJURP and CENP-A become necessary to support rapidly dividing p53 deficient cancer cells. Alterations in CENP-A posttranslational modifications are also linked to chromosome segregation errors and CIN. Collectively, CENP-A is pivotal to genomic stability through centromere maintenance, perturbation of which can lead to tumorigenesis.

scholarly journals Propagation of centromeric chromatin requires exit from mitosis

2007 ◽  
Vol 176 (6) ◽  
pp. 795-805 ◽  
Author(s):  
Lars E.T. Jansen ◽  
Ben E. Black ◽  
Daniel R. Foltz ◽  
Don W. Cleveland
Keyword(s):  
Protein A ◽  
S Phase ◽  
Epigenetic Mark ◽  
Multiprotein Complex ◽  
Centromeric Chromatin ◽  
Microtubule Attachment ◽  
Centromere Protein A ◽  
Histone H3 Variant ◽  
Multiple Cell ◽  
Labeling Approach

Centromeres direct chromosomal inheritance by nucleating assembly of the kinetochore, a large multiprotein complex required for microtubule attachment during mitosis. Centromere identity in humans is epigenetically determined, with no DNA sequence either necessary or sufficient. A prime candidate for the epigenetic mark is assembly into centromeric chromatin of centromere protein A (CENP-A), a histone H3 variant found only at functional centromeres. A new covalent fluorescent pulse-chase labeling approach using SNAP tagging has now been developed and is used to demonstrate that CENP-A bound to a mature centromere is quantitatively and equally partitioned to sister centromeres generated during S phase, thereby remaining stably associated through multiple cell divisions. Loading of nascent CENP-A on the megabase domains of replicated centromere DNA is shown to require passage through mitosis but not microtubule attachment. Very surprisingly, assembly and stabilization of new CENP-A–containing nucleosomes is restricted exclusively to the subsequent G1 phase, demonstrating direct coupling between progression through mitosis and assembly/maturation of the next generation of centromeres.

scholarly journals Three-dimensional localization of CENP-A suggests a complex higher order structure of centromeric chromatin

2008 ◽  
Vol 183 (7) ◽  
pp. 1193-1202 ◽  
Author(s):  
Owen J. Marshall ◽  
Alan T. Marshall ◽  
K.H. Andy Choo
Keyword(s):  
Chromatin Immunoprecipitation ◽  
Protein A ◽  
Three Dimensional ◽  
Higher Order ◽  
Order Structure ◽  
Mitotic Chromosomes ◽  
Centromeric Chromatin ◽  
Centromere Protein A ◽  
Transmission Electron ◽  
Histone H3 Variant

The histone H3 variant centromere protein A (CENP-A) is central to centromere formation throughout eukaryotes. A long-standing question in centromere biology has been the organization of CENP-A at the centromere and its implications for the structure of centromeric chromatin. In this study, we describe the three-dimensional localization of CENP-A at the inner kinetochore plate through serial-section transmission electron microscopy of human mitotic chromosomes. At the kinetochores of normal centromeres and at a neocentromere, CENP-A occupies a compact domain at the inner kinetochore plate, stretching across two thirds of the length of the constriction but encompassing only one third of the constriction width and height. Within this domain, evidence of substructure is apparent. Combined with previous chromatin immunoprecipitation results (Saffery, R., H. Sumer, S. Hassan, L.H. Wong, J.M. Craig, K. Todokoro, M. Anderson, A. Stafford, and K.H.A. Choo. 2003. Mol. Cell. 12:509–516; Chueh, A.C., L.H. Wong, N. Wong, and K.H.A. Choo. 2005. Hum. Mol. Genet. 14:85–93), our data suggest that centromeric chromatin is arranged in a coiled 30-nm fiber that is itself coiled or folded to form a higher order structure.

scholarly journals Functional genomics identifies a Myb domain–containing protein family required for assembly of CENP-A chromatin

2007 ◽  
Vol 176 (6) ◽  
pp. 757-763 ◽  
Author(s):  
Paul S. Maddox ◽  
Francie Hyndman ◽  
Joost Monen ◽  
Karen Oegema ◽  
Arshad Desai
Keyword(s):  
Functional Genomics ◽  
Protein A ◽  
Common Mechanism ◽  
Centromeric Chromatin ◽  
Kinetochore Assembly ◽  
Centromere Protein A ◽  
Close Proximity ◽  
Single Protein ◽  
Histone H3 Variant ◽  
New Protein

Nucleosomes containing the centromere-specific histone H3 variant centromere protein A (CENP-A) create the chromatin foundation for kinetochore assembly. To understand the mechanisms that selectively target CENP-A to centromeres, we took a functional genomics approach in the nematode Caenorhabditis elegans, in which failure to load CENP-A results in a signature kinetochore-null (KNL) phenotype. We identified a single protein, KNL-2, that is specifically required for CENP-A incorporation into chromatin. KNL-2 and CENP-A localize to centromeres throughout the cell cycle in an interdependent manner and coordinately direct chromosome condensation, kinetochore assembly, and chromosome segregation. The isolation of KNL-2–associated chromatin coenriched CENP-A, indicating their close proximity on DNA. KNL-2 defines a new conserved family of Myb DNA-binding domain–containing proteins. The human homologue of KNL-2 is also specifically required for CENP-A loading and kinetochore assembly but is only transiently present at centromeres after mitotic exit. These results implicate a new protein class in the assembly of centromeric chromatin and suggest that holocentric and monocentric chromosomes share a common mechanism for CENP-A loading.

scholarly journals Identification and characterization of centromeric sequences in Xenopus laevis

2020 ◽  
Author(s):  
Owen K Smith ◽  
Charles Limouse ◽  
Kelsey A Fryer ◽  
Nicole A Teran ◽  
Kousik Sundararajan ◽  
...  
Keyword(s):  
Xenopus Laevis ◽  
Repetitive Sequences ◽  
Protein A ◽  
Repeat Sequences ◽  
Centromere Protein A ◽  
Histone H3 Variant ◽  
Egg Extracts ◽  
Genomic Studies

AbstractCentromeres play an essential function in cell division by specifying the site of kinetochore formation on each chromosome for mitotic spindle attachment. Centromeres are defined epigenetically by the histone H3 variant CEntromere Protein A (CENP-A). CENP-A nucleosomes maintain the centromere by designating the site for new CENP-A assembly after dilution by replication. Vertebrate centromeres assemble on tandem arrays of repetitive sequences but the function of repeat DNA in centromere formation has been challenging to dissect due to the difficulty in manipulating centromeres in cells. Xenopus laevis egg extracts assemble centromeres in vitro, providing a system for studying centromeric DNA functions. However, centromeric sequences in X. laevis have not been extensively characterized. In this study we combine CENP-A ChIP-seq with a k-mer based analysis approach to identify the X. laevis centromere repeat sequences. By in situ hybridization we show that X. laevis centromeres contain diverse repeat sequences and we map the centromere position on each X. laevis chromosome using the distribution of centromere enriched k-mers. Our identification of X. laevis centromere sequences enables previously unapproachable centromere genomic studies. Our approach should be broadly applicable for the analysis of centromere and other repetitive sequences in any organism.

scholarly journals Trans-generational inheritance of centromere identity requires the CENP-A N-terminal tail in the C. elegans maternal germ line

2020 ◽  
Author(s):  
Reinier F. Prosée ◽  
Joanna M. Wenda ◽  
Caroline Gabus ◽  
Kamila Delaney ◽  
Francoise Schwager ◽  
...  
Keyword(s):  
De Novo ◽  
Germ Line ◽  
Protein A ◽  
Histone H3 ◽  
Centromeric Chromatin ◽  
C Elegans ◽  
Centromere Function ◽  
Centromere Protein A ◽  
Histone H3 Variant ◽  
Meiosis I

AbstractCentromere protein A (CENP-A) is a histone H3 variant that defines centromeric chromatin and is essential for centromere function. In most eukaryotes CENP-A-containing chromatin is epigenetically maintained, and centromere identity is inherited from one cell cycle to the next. In the germ line of the holocentric nematode Caenorhabditis elegans, this inheritance cycle is disrupted. CENP-A is removed at the mitosis-to-meiosis transition and is established de novo on chromatin during diplotene of meiosis I. Here we show that the N-terminal tail of CENP-A is required for the de novo establishment of centromeres, but dispensable for centromere maintenance during embryogenesis. Worms homozygous for a CENP-A tail deletion maintain a functional centromere during development, but give rise to inviable offspring because they fail to re-establish centromeres in the maternal germ line. We identify the N-terminal tail of CENP-A as a critical domain for the interaction with the conserved kinetochore protein KNL-2, and argue that this interaction plays an important role in setting centromere identity in the germ line. We conclude that centromere establishment and maintenance are functionally distinct in C. elegans.

scholarly journals CENP-C recruits M18BP1 to centromeres to promote CENP-A chromatin assembly

2011 ◽  
Vol 194 (6) ◽  
pp. 855-871 ◽  
Author(s):  
Ben Moree ◽  
Corey B. Meyer ◽  
Colin J. Fuller ◽  
Aaron F. Straight
Keyword(s):  
Chromosome Segregation ◽  
Protein A ◽  
Histone H3 ◽  
Chromatin Assembly ◽  
Nucleosome Assembly ◽  
Chromosomal Locus ◽  
C Protein ◽  
Centromere Protein A ◽  
Histone H3 Variant ◽  
Complex Protein

Eukaryotic chromosomes segregate by attaching to microtubules of the mitotic spindle through a chromosomal microtubule binding site called the kinetochore. Kinetochores assemble on a specialized chromosomal locus termed the centromere, which is characterized by the replacement of histone H3 in centromeric nucleosomes with the essential histone H3 variant CENP-A (centromere protein A). Understanding how CENP-A chromatin is assembled and maintained is central to understanding chromosome segregation mechanisms. CENP-A nucleosome assembly requires the Mis18 complex and the CENP-A chaperone HJURP. These factors localize to centromeres in telophase/G1, when new CENP-A chromatin is assembled. The mechanisms that control their targeting are unknown. In this paper, we identify a mechanism for recruiting the Mis18 complex protein M18BP1 to centromeres. We show that depletion of CENP-C prevents M18BP1 targeting to metaphase centromeres and inhibits CENP-A chromatin assembly. We find that M18BP1 directly binds CENP-C through conserved domains in the CENP-C protein. Thus, CENP-C provides a link between existing CENP-A chromatin and the proteins required for new CENP-A nucleosome assembly.

scholarly journals Constitutive centromere-associated network contacts confer differential stability on CENP-A nucleosomes in vitro and in the cell

2018 ◽  
Vol 29 (6) ◽  
pp. 751-762 ◽  
Author(s):  
Shengya Cao ◽  
Keda Zhou ◽  
Zhening Zhang ◽  
Karolin Luger ◽  
Aaron F. Straight
Keyword(s):  
Protein A ◽  
Histone H3 ◽  
Intrinsic Property ◽  
Cell Cycles ◽  
Centromere Protein A ◽  
Histone H3 Variant ◽  
Differential Stability ◽  
The Stability

Eukaryotic centromeres are defined by the presence of nucleosomes containing the histone H3 variant, centromere protein A (CENP-A). Once incorporated at centromeres, CENP-A nucleosomes are remarkably stable, exhibiting no detectable loss or exchange over many cell cycles. It is currently unclear whether this stability is an intrinsic property of CENP-A containing chromatin or whether it arises from proteins that specifically associate with CENP-A chromatin. Two proteins, CENP-C and CENP-N, are known to bind CENP-A human nucleosomes directly. Here we test the hypothesis that CENP-C or CENP-N stabilize CENP-A nucleosomes in vitro and in living cells. We show that CENP-N stabilizes CENP-A nucleosomes alone and additively with CENP-C in vitro. However, removal of CENP-C and CENP-N from cells, or mutating CENP-A so that it no longer interacts with CENP-C or CENP-N, had no effect on centromeric CENP-A stability in vivo. Thus, the stability of CENP-A nucleosomes in chromatin does not arise solely from its interactions with CENP-C or CENP-N.

How to build a centromere: from centromeric and pericentromeric chromatin to kinetochore assemblyThis paper is one of a selection of papers published in this Special Issue, entitled 27th International West Coast Chromatin and Chromosome Conference, and has undergone the Journal's usual peer review process.

2006 ◽  
Vol 84 (4) ◽  
pp. 619-639 ◽  
Author(s):  
Larissa J. Vos ◽  
Jakub K. Famulski ◽  
Gordon K.T. Chan
Keyword(s):  
Protein Interactions ◽  
Core Protein ◽  
Peer Review Process ◽  
Protein A ◽  
Histone Deacetylation ◽  
Protein Protein Interactions ◽  
Centromere Protein A ◽  
Histone H3 Variant ◽  
Base Excision ◽  
Specialized Region

The assembly of the centromere, a specialized region of DNA along with a constitutive protein complex which resides at the primary constriction and is the site of kinetochore formation, has been puzzling biologists for many years. Recent advances in the fields of chromatin, microscopy, and proteomics have shed a new light on this complex and essential process. Here we review recently discovered mechanisms and proteins involved in determining mammalian centromere location and assembly. The centromeric core protein CENP-A, a histone H3 variant, is hypothesized to designate centromere localization by incorporation into centromere-specific nucleosomes and is essential for the formation of a functional kinetochore. It has been found that centromere localization of centromere protein A (CENP-A), and therefore centromere determination, requires proteins involved in histone deacetylation, as well as base excision DNA repair pathways and proteolysis. In addition to the incorporation of CENP-A at the centromere, the formation of heterochromatin through histone methylation and RNA interference is also crucial for centromere formation. The assembly of the centromere and kinetochore is complex and interdependent, involving epigenetics and hierarchical protein–protein interactions.

scholarly journals Centromeric chromatin gets loaded

2007 ◽  
Vol 176 (6) ◽  
pp. 735-736 ◽  
Author(s):  
Christopher W. Carroll ◽  
Aaron F. Straight
Keyword(s):  
Chromosome Segregation ◽  
Molecular Mechanisms ◽  
Protein A ◽  
Histone H3 ◽  
Chromatin Assembly ◽  
Centromeric Chromatin ◽  
Kinetochore Assembly ◽  
Centromere Protein A ◽  
Specific Loading ◽  
Histone H3 Variant

Centromeric nucleosomes contain a histone H3 variant called centromere protein A (CENP-A) that is required for kinetochore assembly and chromosome segregation. Two new studies, Jansen et al. (see p. 795 of this issue) and Maddox et al. (see p. 757 of this issue), address when CENP-A is deposited at centromeres during the cell division cycle and identify an evolutionally conserved protein required for CENP-A deposition. Together, these studies advance our understanding of centromeric chromatin assembly and provide a framework for investigating the molecular mechanisms that underlie the centromere-specific loading of CENP-A.

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