New insights into the helical structure of 30-nm chromatin fibers.
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ABSTRACT:
SUBMITTER: Chen P
PROVIDER: S-EPMC4085281 | biostudies-literature | 2014 Jul
REPOSITORIES: biostudies-literature
Publications
New insights into the helical structure of 30-nm chromatin fibers.
Chen Ping P Zhu Ping P Li Guohong G
Protein & cell 20140701 7
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Project description:Chromatin folding in eukaryotes fits the genome into the limited volume of the cell nucleus. Formation of higher-order chromatin structures attenuates DNA accessibility, thus contributing to the control of essential genome functions such as transcription, DNA replication, and repair. The 30-nm fiber is thought to be the first hierarchical level of chromatin folding, but the nucleosome arrangement in the compact 30-nm fiber was previously unknown. We used cryoelectron tomography of vitreous sections to determine the structure of the compact, native 30-nm fiber of avian erythrocyte nuclei. The predominant geometry of the 30-nm fiber revealed by subtomogram averaging is a left-handed two-start helix with approximately 6.5 nucleosomes per 11 nm, in which the nucleosomes are juxtaposed face-to-face but are shifted off their superhelical axes with an axial translation of approximately 3.4 nm and an azimuthal rotation of approximately 54°. The nucleosomes produce a checkerboard pattern when observed in the direction perpendicular to the fiber axis but are not interdigitated. The nucleosome packing within the fibers shows larger center-to-center internucleosomal distances than previously anticipated, thus excluding the possibility of core-to-core interactions, explaining how transcription and regulation factors can access nucleosomes.
| S-EPMC3193215 | biostudies-other
Project description:The structure of compact 30-nm chromatin fibres is still debated. We present here a novel unified model that reconciles all experimental observations into a single framework. We propose that compact fibres are formed by the interdigitation of the two nucleosome stacks in a 2-start crossed-linker structure to form a single stack. This process requires that the dyad orientation of successive nucleosomes relative to the helical axis alternates. The model predicts that, as observed experimentally, the fibre-packing density should increase in a stepwise manner with increasing linker length. This model structure can also incorporate linker DNA of varying lengths.
| S-EPMC4863496 | biostudies-literature
Project description:Why most of the in vivo experiments do not find the 30-nm chromatin fiber, well studied in vitro, is a puzzle. Two basic physical inputs that are crucial for understanding the structure of the 30-nm fiber are the stiffness of the linker DNA and the relative orientations of the DNA entering/exiting nucleosomes. Based on these inputs we simulate chromatin structure and show that the presence of non-histone proteins, which bind and locally bend linker DNA, destroys any regular higher order structures (e.g., zig-zag). Accounting for the bending geometry of proteins like nhp6 and HMG-B, our theory predicts phase-diagram for the chromatin structure as a function of DNA-bending non-histone protein density and mean linker DNA length. For a wide range of linker lengths, we show that as we vary one parameter, that is, the fraction of bent linker region due to non-histone proteins, the steady-state structure will show a transition from zig-zag to an irregular structure-a structure that is reminiscent of what is observed in experiments recently. Our theory can explain the recent in vivo observation of irregular chromatin having co-existence of finite fraction of the next-neighbor (i + 2) and neighbor (i + 1) nucleosome interactions.
| S-EPMC5305278 | biostudies-literature
Project description:The structure of the 30-nm chromatin fibre is an important determinant of the regulation of eukaryotic transcription. A fundamental issue is whether the stacking of nucleosomes in this fibre is organized as a one-start or two-start helix. We argue that all recent experimental data are compatible with a two-start helix and propose that the topology of the fibre, but not the mode of stacking the nucleosomes, is dependent on the length of the linker DNA. This arrangement conserves nucleosome stacking and thus the external morphology of the fibre, and also ensures that the fibre adopts the highest available packing density.
| S-EPMC2267236 | biostudies-literature
Project description:How a long strand of genomic DNA is compacted into a mitotic chromosome remains one of the basic questions in biology. The nucleosome fibre, in which DNA is wrapped around core histones, has long been assumed to be folded into a 30-nm chromatin fibre and further hierarchical regular structures to form mitotic chromosomes, although the actual existence of these regular structures is controversial. Here, we show that human mitotic HeLa chromosomes are mainly composed of irregularly folded nucleosome fibres rather than 30-nm chromatin fibres. Our comprehensive and quantitative study using cryo-electron microscopy and synchrotron X-ray scattering resolved the long-standing contradictions regarding the existence of 30-nm chromatin structures and detected no regular structure >11 nm. Our finding suggests that the mitotic chromosome consists of irregularly arranged nucleosome fibres, with a fractal nature, which permits a more dynamic and flexible genome organization than would be allowed by static regular structures.
| S-EPMC3321210 | biostudies-literature
Project description:Molecular dynamics (MD) simulations based on the implicit solvent generalized Born (GB) models can provide significant computational advantages over the traditional explicit solvent simulations. However, the standard GB becomes prohibitively expensive for all-atom simulations of large structures; the model scales poorly, ∼n2, with the number of solute atoms. Here we combine our recently developed optimal point charge approximation (OPCA) with the hierarchical charge partitioning (HCP) approximation to present an ∼n log n multiscale, yet fully atomistic, GB model (GB-HCPO). The HCP approximation exploits the natural organization of biomolecules (atoms, groups, chains, and complexes) to partition the structure into multiple hierarchical levels of components. OPCA approximates the charge distribution for each of these components by a small number of point charges so that the low order multipole moments of these components are optimally reproduced. The approximate charges are then used for computing electrostatic interactions with distant components, while the full set of atomic charges are used for nearby components. We show that GB-HCPO can deliver up to 2 orders of magnitude speedup compared to the standard GB, with minimal impact on its accuracy. For large structures, GB-HCPO can approach the same nominal speed, as in nanoseconds per day, as the highly optimized explicit-solvent simulation based on particle mesh Ewald (PME). The increase in the nominal simulation speed, relative to the standard GB, coupled with substantially faster sampling of conformational space, relative to the explicit solvent, makes GB-HCPO a suitable candidate for MD simulation of large atomistic systems in implicit solvent. As a practical demonstration, we use GB-HCPO simulation to refine a ∼1.16 million atom structure of 30 nm chromatin fiber (40 nucleosomes). The refined structure suggests important details about spatial organization of the linker DNA and the histone tails in the fiber: (1) the linker DNA fills the core region, allowing the H3 histone tails to interact with the linker DNA, which is consistent with experiment; (2) H3 and H4 tails are found mostly in the core of the structure, closer to the helical axis of the fiber, while H2A and H2B are mostly solvent exposed. Potential functional consequences of these findings are discussed. GB-HCPO is implemented in the open source MD software NAB in Amber 2016.
| S-EPMC5649046 | biostudies-literature
Project description:Although the formation of 30-nm chromatin fibers is thought to be the most basic event of chromatin compaction, it remains controversial because high-resolution imaging of chromatin in living eukaryotic cells had not been possible until now. Cryo-electron microscopy of vitreous sections is a relatively new technique, which enables direct high-resolution observation of the cell structures in a close-to-native state. We used cryo-electron microscopy and image processing to further investigate the presence of 30-nm chromatin fibers in human mitotic chromosomes. HeLa S3 cells were vitrified by high-pressure freezing, thin-sectioned, and then imaged under the cryo-electron microscope without any further chemical treatment or staining. For an unambiguous interpretation of the images, the effects of the contrast transfer function were computationally corrected. The mitotic chromosomes of the HeLa S3 cells appeared as compact structures with a homogeneous grainy texture, in which there were no visible 30-nm fibers. Power spectra of the chromosome images also gave no indication of 30-nm chromatin folding. These results, together with our observations of the effects of chromosome swelling, strongly suggest that, within the bulk of compact metaphase chromosomes, the nucleosomal fiber does not undergo 30-nm folding, but exists in a highly disordered and interdigitated state, which is, on the local scale, comparable with a polymer melt.
| S-EPMC2604964 | biostudies-literature
Project description:The existence of a 30-nm fiber as a basic folding unit for DNA packaging has remained a topic of active discussion. Here, we characterize the supramolecular structures formed by reversible Mg(2+)-dependent self-association of linear 12-mer nucleosomal arrays using microscopy and physicochemical approaches. These reconstituted chromatin structures, which we call "oligomers", are globular throughout all stages of cooperative assembly and range in size from ~50 nm to a maximum diameter of ~1,000 nm. The nucleosomal arrays were packaged within the oligomers as interdigitated 10-nm fibers, rather than folded 30-nm structures. Linker DNA was freely accessible to micrococcal nuclease, although the oligomers remained partially intact after linker DNA digestion. The organization of chromosomal fibers in human nuclei in situ was stabilized by 1 mM MgCl2, but became disrupted in the absence of MgCl2, conditions that also dissociated the oligomers in vitro These results indicate that a 10-nm array of nucleosomes has the intrinsic ability to self-assemble into large chromatin globules stabilized by nucleosome-nucleosome interactions, and suggest that the oligomers are a good in vitro model for investigating the structure and organization of interphase chromosomes.
| S-EPMC4868957 | biostudies-literature
Project description:Chromatin structure plays a fundamental role in the regulation of nuclear processes such as DNA transcription, replication, recombination, and repair. Despite considerable efforts during three decades, the structure of the 30-nm chromatin fiber remains controversial. To define fiber dimensions accurately, we have produced very long and regularly folded 30-nm fibers from in vitro reconstituted nucleosome arrays containing the linker histone and with increasing nucleosome repeat lengths (10 to 70 bp of linker DNA). EM measurements show that the dimensions of these fully folded fibers do not increase linearly with increasing linker length, a finding that is inconsistent with two-start helix models. Instead, we find that there are two distinct classes of fiber structure, both with unexpectedly high nucleosome density: arrays with 10 to 40 bp of linker DNA all produce fibers with a diameter of 33 nm and 11 nucleosomes per 11 nm, whereas arrays with 50 to 70 bp of linker DNA all produce 44-nm-wide fibers with 15 nucleosomes per 11 nm. Using the physical constraints imposed by these measurements, we have built a model in which tight nucleosome packing is achieved through the interdigitation of nucleosomes from adjacent helical gyres. Importantly, the model closely matches raw image projections of folded chromatin arrays recorded in the solution state by using electron cryo-microscopy.
| S-EPMC1436021 | biostudies-literature
Project description:The mechanism by which chromatin is decondensed to permit access to DNA is largely unknown. Here, using a model nucleosome array reconstituted from recombinant histone octamers, we have defined the relative contribution of the individual histone octamer N-terminal tails as well as the effect of a targeted histone tail acetylation on the compaction state of the 30 nm chromatin fiber. This study goes beyond previous studies as it is based on a nucleosome array that is very long (61 nucleosomes) and contains a stoichiometric concentration of bound linker histone, which is essential for the formation of the 30 nm chromatin fiber. We find that compaction is regulated in two steps: Introduction of H4 acetylated to 30% on K16 inhibits compaction to a greater degree than deletion of the H4 N-terminal tail. Further decompaction is achieved by removal of the linker histone.
| S-EPMC3870898 | biostudies-literature