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The cryo-EM structure of the CENP-A nucleosome in complex with the phosphorylated CENP-C:: CENP-A nucleosome in complex with CENP-C motif (655-675) and CENP-N N-terminal domain (1-211)

ABSTRACT:

SUBMITTER: Fukagawa Tatsuo

PROVIDER: EMPIAR-10571 | biostudies-other |

REPOSITORIES: biostudies-other

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Cryo-EM structure of the CENP-A nucleosome in complex with phosphorylated CENP-C.

Ariyoshi Mariko M Makino Fumiaki F Watanabe Reito R Nakagawa Reiko R Kato Takayuki T Namba Keiichi K Arimura Yasuhiro Y Fujita Risa R Kurumizaka Hitoshi H Okumura Ei-Ichi EI Hara Masatoshi M Fukagawa Tatsuo T

The EMBO journal 20210119 5

The CENP-A nucleosome is a key structure for kinetochore assembly. Once the CENP-A nucleosome is established in the centromere, additional proteins recognize the CENP-A nucleosome to form a kinetochore. CENP-C and CENP-N are CENP-A binding proteins. We previously demonstrated that vertebrate CENP-C binding to the CENP-A nucleosome is regulated by CDK1-mediated CENP-C phosphorylation. However, it is still unknown how the phosphorylation of CENP-C regulates its binding to CENP-A. It is also not co ...[more]

PMID: 33463726

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The cryo-EM structure of the CENP-A nucleosome in complex with the phosphorylated CENP-C: CENP-A nucleosome in complex with phosphorylated CENP-C C-terminal domain (601-864) and CENP-N N-terminal domain (1-211)

Project description: Not available

| EMPIAR-10565 | biostudies-other

The cryo-EM structure of the CENP-A nucleosome in complex with the phosphorylated CENP-C:: CENP-A nucleosome in complex with phosphorylated CENP-C C-terminal domain(601-864)

Project description: Not available

| EMPIAR-10569 | biostudies-other

Cryo-EM structure of SWI/SNF complex bound to a nucleosome.

Project description:The chromatin-remodelling complex SWI/SNF is highly conserved and has critical roles in various cellular processes, including transcription and DNA-damage repair1,2. It hydrolyses ATP to remodel chromatin structure by sliding and evicting histone octamers3-8, creating DNA regions that become accessible to other essential factors. However, our mechanistic understanding of the remodelling activity is hindered by the lack of a high-resolution structure of complexes from this family. Here we report the cryo-electron microscopy structure of Saccharomyces cerevisiae SWI/SNF bound to a nucleosome, at near-atomic resolution. In the structure, the actin-related protein (Arp) module is sandwiched between the ATPase and the rest of the complex, with the Snf2 helicase-SANT associated (HSA) domain connecting all modules. The body contains an assembly scaffold composed of conserved subunits Snf12 (also known as SMARCD or BAF60), Snf5 (also known as SMARCB1, BAF47 or INI1) and an asymmetric dimer of Swi3 (also known as SMARCC, BAF155 or BAF170). Another conserved subunit, Swi1 (also known as ARID1 or BAF250), resides in the core of SWI/SNF, acting as a molecular hub. We also observed interactions between Snf5 and the histones at the acidic patch, which could serve as an anchor during active DNA translocation. Our structure enables us to map and rationalize a subset of cancer-related mutations in the human SWI/SNF complex and to propose a model for how SWI/SNF recognizes and remodels the +1 nucleosome to generate nucleosome-depleted regions during gene activation9.

| S-EPMC7319049 | biostudies-literature

Atomic resolution cryo-EM structure of a native-like CENP-A nucleosome aided by an antibody fragment.

Project description:Genomic DNA in eukaryotes is organized into chromatin through association with core histones to form nucleosomes, each distinguished by their DNA sequences and histone variants. Here, we used a single-chain antibody fragment (scFv) derived from the anti-nucleosome antibody mAb PL2-6 to stabilize human CENP-A nucleosome containing a native α-satellite DNA and solved its structure by the cryo-electron microscopy (cryo-EM) to 2.6 Å resolution. In comparison, the corresponding cryo-EM structure of the free CENP-A nucleosome could only reach 3.4 Å resolution. We find that scFv binds to a conserved acidic patch on the histone H2A-H2B dimer without perturbing the nucleosome structure. Our results provide an atomic resolution cryo-EM structure of a nucleosome and insight into the structure and function of the CENP-A nucleosome. The scFv approach is applicable to the structural determination of other native-like nucleosomes with distinct DNA sequences.

| S-EPMC6534667 | biostudies-literature

Cryo-EM structure of the human MLL1 core complex bound to the nucleosome.

Project description:Mixed lineage leukemia (MLL) family histone methyltransferases are enzymes that deposit histone H3 Lys4 (K4) mono-/di-/tri-methylation and regulate gene expression in mammals. Despite extensive structural and biochemical studies, the molecular mechanisms whereby the MLL complexes recognize histone H3K4 within nucleosome core particles (NCPs) remain unclear. Here we report the single-particle cryo-electron microscopy (cryo-EM) structure of the NCP-bound human MLL1 core complex. We show that the MLL1 core complex anchors to the NCP via the conserved RbBP5 and ASH2L, which interact extensively with nucleosomal DNA and the surface close to the N-terminal tail of histone H4. Concurrent interactions of RbBP5 and ASH2L with the NCP uniquely align the catalytic MLL1SET domain at the nucleosome dyad, thereby facilitating symmetrical access to both H3K4 substrates within the NCP. Our study sheds light on how the MLL1 complex engages chromatin and how chromatin binding promotes MLL1 tri-methylation activity.

| S-EPMC6895043 | biostudies-literature

Cryo-EM structure of the human Sirtuin 6-nucleosome complex

Project description: Not available

| EMPIAR-11427 | biostudies-other

Phase-plate cryo-EM structure of the Widom 601 CENP-A nucleosome core particle reveals differential flexibility of the DNA ends.

Project description:The histone H3 variant CENP-A marks centromeres epigenetically and is essential for mitotic fidelity. Previous crystallographic studies of the CENP-A nucleosome core particle (NCP) reconstituted with a human ?-satellite DNA derivative revealed both DNA ends to be highly flexible, a feature important for CENP-A mitotic functions. However, recent cryo-EM studies of CENP-A NCP complexes comprising primarily Widom 601 DNA reported well-ordered DNA ends. Here, we report the cryo-EM structure of the CENP-A 601 NCP determined by Volta phase-plate imaging. The data reveal that one ('left') 601 DNA end is well ordered whereas the other ('right') end is flexible and partly detached from the histone core, suggesting sequence-dependent dynamics of the DNA termini. Indeed, a molecular dynamics simulation of the CENP-A 601 NCP confirmed the distinct dynamics of the two DNA extremities. Reprocessing the image data using two-fold symmetry yielded a cryo-EM map in which both DNA ends appeared well ordered, indicating that such an artefact may inadvertently arise if NCP asymmetry is lost during image processing. These findings enhance our understanding of the dynamic features that discriminate CENP-A from H3 nucleosomes by revealing that DNA end flexibility can be fine-tuned in a sequence-dependent manner.

| S-EPMC7261176 | biostudies-literature

Cryo-EM of nucleosome core particle interactions in trans.

Project description:Nucleosomes, the basic unit of chromatin, are repetitively spaced along DNA and regulate genome expression and maintenance. The long linear chromatin molecule is extensively condensed to fit DNA inside the nucleus. How distant nucleosomes interact to build tertiary chromatin structure remains elusive. In this study, we used cryo-EM to structurally characterize different states of long range nucleosome core particle (NCP) interactions. Our structures show that NCP pairs can adopt multiple conformations, but, commonly, two NCPs are oriented with the histone octamers facing each other. In this conformation, the dyad of both nucleosome core particles is facing the same direction, however, the NCPs are laterally shifted and tilted. The histone octamer surface and histone tails in trans NCP pairs remain accessible to regulatory proteins. The overall conformational flexibility of the NCP pair suggests that chromatin tertiary structure is dynamic and allows access of various chromatin modifying machineries to nucleosomes.

| S-EPMC5935684 | biostudies-literature

Cryo-EM structures of remodeler-nucleosome intermediates suggest allosteric control through the nucleosome.

Project description:The SNF2h remodeler slides nucleosomes most efficiently as a dimer, yet how the two protomers avoid a tug-of-war is unclear. Furthermore, SNF2h couples histone octamer deformation to nucleosome sliding, but the underlying structural basis remains unknown. Here we present cryo-EM structures of SNF2h-nucleosome complexes with ADP-BeFx that capture two potential reaction intermediates. In one structure, histone residues near the dyad and in the H2A-H2B acidic patch, distal to the active SNF2h protomer, appear disordered. The disordered acidic patch is expected to inhibit the second SNF2h protomer, while disorder near the dyad is expected to promote DNA translocation. The other structure doesn't show octamer deformation, but surprisingly shows a 2 bp translocation. FRET studies indicate that ADP-BeFx predisposes SNF2h-nucleosome complexes for an elemental translocation step. We propose a model for allosteric control through the nucleosome, where one SNF2h protomer promotes asymmetric octamer deformation to inhibit the second protomer, while stimulating directional DNA translocation.

| S-EPMC6611695 | biostudies-literature

CENP-C unwraps the human CENP-A nucleosome through the H2A C-terminal tail.

Project description:Centromeres are defined epigenetically by nucleosomes containing the histone H3 variant CENP-A, upon which the constitutive centromere-associated network of proteins (CCAN) is built. CENP-C is considered to be a central organizer of the CCAN. We provide new molecular insights into the structure of human CENP-A nucleosomes, in isolation and in complex with the CENP-C central region (CENP-CCR ), the main CENP-A binding module of human CENP-C. We establish that the short ?N helix of CENP-A promotes DNA flexibility at the nucleosome ends, independently of the sequence it wraps. Furthermore, we show that, in vitro, two regions of human CENP-C (CENP-CCR and CENP-Cmotif ) both bind exclusively to the CENP-A nucleosome. We find CENP-CCR to bind with high affinity due to an extended hydrophobic area made up of CENP-AV 532 and CENP-AV 533 . Importantly, we identify two key conformational changes within the CENP-A nucleosome upon CENP-C binding. First, the loose DNA wrapping of CENP-A nucleosomes is further exacerbated, through destabilization of the H2A C-terminal tail. Second, CENP-CCR rigidifies the N-terminal tail of H4 in the conformation favoring H4K20 monomethylation, essential for a functional centromere.

| S-EPMC6776904 | biostudies-literature

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