Nonlinear mechanics of lamin filaments and the meshwork topology build an emergent nuclear lamina

AbstractThe nuclear lamina—a meshwork of intermediate filaments termed lamins—is primarily responsible for the mechanical stability of the nucleus in multicellular organisms. However, structural-mechanical characterization of lamin filaments assembled in situ remains elusive. Here, we apply an integrative approach combining atomic force microscopy, cryo-electron tomography, network analysis, and molecular dynamics simulations to directly measure the mechanical response of single lamin filaments in three-dimensional meshwork. Endogenous lamin filaments portray non-Hookean behavior – they deform reversibly at a few hundred picoNewtons and stiffen at nanoNewton forces. The filaments are extensible, strong and tough similar to natural silk and superior to the synthetic polymer Kevlar®. Graph theory analysis shows that the lamin meshwork is not a random arrangement of filaments but exhibits small-world properties. Our results suggest that lamin filaments arrange to form an emergent meshwork whose topology dictates the mechanical properties of individual filaments. The quantitative insights imply a role of meshwork topology in laminopathies.

Download Full-text

Nonlinear mechanics of lamin filaments and the meshwork topology build an emergent nuclear lamina

10.1101/846550 ◽

2019 ◽

Author(s):

K. Tanuj Sapra ◽

Zhao Qin ◽

Anna Dubrovsky-Gaupp ◽

Ueli Aebi ◽

Daniel J. Müller ◽

...

Keyword(s):

Mechanical Response ◽

Mechanical Stability ◽

Electron Tomography ◽

Three Dimensional ◽

Small World ◽

Nuclear Lamina ◽

Integrative Approach ◽

Graph Theory Analysis ◽

Dynamics Simulations

AbstractThe nuclear lamina – a meshwork of intermediate filaments termed lamins – functions as a mechanotransduction interface between the extracellular matrix and the nucleus via the cytoskeleton. Although lamins are primarily responsible for the mechanical stability of the nucleus in multicellular organisms, in situ characterization of lamin filaments under tension has remained elusive. Here, we apply an integrative approach combining atomic force microscopy, cryo-electron tomography, network analysis, and molecular dynamics simulations to directly measure the mechanical response of single lamin filaments in its three-dimensional meshwork. Endogenous lamin filaments portray non-Hookean behavior – they deform reversibly under a force of a few hundred picoNewtons and stiffen at nanoNewton forces. The filaments are extensible, strong and tough, similar to natural silk and superior to the synthetic polymer Kevlar®. Graph theory analysis shows that the lamin meshwork is not a random arrangement of filaments but the meshwork topology follows ‘small world’ properties. Our results suggest that the lamin filaments arrange to form a robust, emergent meshwork that dictates the mechanical properties of individual lamin filaments. The combined approach provides quantitative insights into the structure-function organization of lamins in situ, and implies a role of meshwork topology in laminopathies.

Download Full-text

Study of Pseudomonas aeruginosa PPF-1 biofilm formation using microfluidics: effect of magnesium on biofilm detachment in engineered conduits

10.33774/chemrxiv-2021-w487v ◽

2021 ◽

Author(s):

William Y. Harvey ◽

Cynthia Gagné-Thivierge ◽

Sepideh Fakari ◽

Jean Barbeau ◽

Steve Charette ◽

...

Keyword(s):

Pseudomonas Aeruginosa ◽

Reference Strain ◽

Mechanical Stability ◽

Three Dimensional ◽

Opportunistic Pathogen ◽

Shear Stresses ◽

Density Maps ◽

Biofilm Detachment ◽

Flow Systems ◽

Dynamics Simulations

The bacterium Pseudomonas aeruginosa is an opportunistic pathogen in certain organisms, including humans, but can also survive and proliferate in natural and engineered water systems. Microfluidic technology can address hydrodynamic questions related to bacterial contamination of water flow systems and infrastructure. In this work, a microfluidic approach was devised to study the effect of shear stresses on biofilms from a dental unit waterline (DUWL)-isolated P. aeruginosa strain, PPF-1. During application of relevant shear stress levels to DUWLs, the response of the PPF-1 biofilm was observed and compared to a clinical P. aeruginosa reference strain, PAO1. The response measurements were repeated for biofilms exposed to additional Mg2+ ions. Using a microfluidic approach to transforming optical density maps into three-dimensional images, we applied computational fluid dynamics simulations and determined the critical shear stresses for biofilm sloughing. In the absence of Mg2+, PPF-1 biofilms showed weaker attachment than PAO1 biofilms, resulting in continuous slough/regrowth cycles triggered by applied shear stresses of 1.42 +/- 0.32 Pa. Introducing Mg2+ into the PPF-1 biofilm culture medium seemed to place the biofilm into a viscoplastic mechanical state, thereby increasing mechanical stability, which resulted in elevated tolerances to shear stresses up to a critical value of 5.43 +/- 1.52 Pa. This resulted in a propensity for less frequent but more catastrophic sloughing events like that observed for the PAO1 reference strain. This suggests that in a low ionic environment, biofilms from the PPF-1 strain can result in higher and more continuous ejection of biofilm materials, possibly leading to increased downstream colonization of engineered flow systems.

Download Full-text

Probing the transition from dislocation jamming to pinning by machine learning

Materials Theory ◽

10.1186/s41313-020-00022-0 ◽

2020 ◽

Vol 4 (1) ◽

Author(s):

Henri Salmenjoki ◽

Lasse Laurson ◽

Mikko J. Alava

Keyword(s):

Machine Learning ◽

Mechanical Response ◽

Collective Motion ◽

Three Dimensional ◽

Dislocation Dynamics ◽

Form Complex ◽

Discrete Dislocation Dynamics ◽

Discrete Dislocation ◽

Two Phases ◽

Dynamics Simulations

Abstract Collective motion of dislocations is governed by the obstacles they encounter. In pure crystals, dislocations form complex structures as they become jammed by their anisotropic shear stress fields. On the other hand, introducing disorder to the crystal causes dislocations to pin to these impeding elements and, thus, leads to a competition between dislocation-dislocation and dislocation-disorder interactions. Previous studies have shown that, depending on the dominating interaction, the mechanical response and the way the crystal yields change.Here we employ three-dimensional discrete dislocation dynamics simulations with varying density of fully coherent precipitates to study this phase transition − from jamming to pinning − using unsupervised machine learning. By constructing descriptors characterizing the evolving dislocation configurations during constant loading, a confusion algorithm is shown to be able to distinguish the systems into two separate phases. These phases agree well with the observed changes in the relaxation rate during the loading. Our results also give insights on the structure of the dislocation networks in the two phases.

Download Full-text

3D Structural Fluctuation of IgG1 Antibody Revealed by Individual Particle Electron Tomography

Scientific Reports ◽

10.1038/srep09803 ◽

2015 ◽

Vol 5 (1) ◽

Cited By ~ 53

Author(s):

Xing Zhang ◽

Lei Zhang ◽

Huimin Tong ◽

Bo Peng ◽

Matthew J. Rames ◽

...

Keyword(s):

Protein Dynamics ◽

Electron Tomography ◽

3D Structure ◽

Three Dimensional ◽

Individual Particle ◽

Single Particle Reconstruction ◽

X Ray Crystallography ◽

Igg1 Antibody ◽

Structural Fluctuation ◽

Dynamics Simulations

Abstract Commonly used methods for determining protein structure, including X-ray crystallography and single-particle reconstruction, often provide a single and unique three-dimensional (3D) structure. However, in these methods, the protein dynamics and flexibility/fluctuation remain mostly unknown. Here, we utilized advances in electron tomography (ET) to study the antibody flexibility and fluctuation through structural determination of individual antibody particles rather than averaging multiple antibody particles together. Through individual-particle electron tomography (IPET) 3D reconstruction from negatively-stained ET images, we obtained 120 ab-initio 3D density maps at an intermediate resolution (~1–3 nm) from 120 individual IgG1 antibody particles. Using these maps as a constraint, we derived 120 conformations of the antibody via structural flexible docking of the crystal structure to these maps by targeted molecular dynamics simulations. Statistical analysis of the various conformations disclosed the antibody 3D conformational flexibility through the distribution of its domain distances and orientations. This blueprint approach, if extended to other flexible proteins, may serve as a useful methodology towards understanding protein dynamics and functions.

Download Full-text

Complete structure of the core signalling unit of the E. coli chemosensory array in an optimised minicell strain

10.1101/723692 ◽

2019 ◽

Author(s):

Alister Burt ◽

C. Keith Cassidy ◽

Peter Ames ◽

Maria Bacia-Verloop ◽

Megghane Baulard ◽

...

Keyword(s):

Electron Tomography ◽

Three Dimensional ◽

Structural Basis ◽

Transmembrane Receptors ◽

Structural Characterisation ◽

E Coli ◽

Chemical Gradients ◽

Membrane Bound ◽

Subtomogram Averaging ◽

Dynamics Simulations

Motile bacteria sense chemical gradients with transmembrane receptors organised in supramolecular signalling arrays.1,2 Understanding stimulus detection and transmission at the molecular level requires precise structural characterisation of the array building block known as a core signalling unit (CSU). Here we introduce a novel E. coli strain that forms small minicells possessing extended and highly ordered chemosensory arrays. We provide a three-dimensional (3D) map of a complete CSU at ~16 Å resolution by cryo-electron tomography (cryo-ET) and subtomogram averaging. This map, combined with previously determined high resolution structures and molecular dynamics simulations, yields an atomistic model of the membrane-bound CSU and enables spatial localisation of its signalling domains. Our work thus offers a solid structural basis for interpretation of existing data and design of new experiments to elucidate signalling mechanisms within the CSU and larger array.

Download Full-text

Grain Size Strengthening in Microcrystalline Copper: A Three-Dimensional Dislocation Dynamics Simulation

Key Engineering Materials ◽

10.4028/www.scientific.net/kem.423.25 ◽

2009 ◽

Vol 423 ◽

pp. 25-32 ◽

Cited By ~ 15

Author(s):

Christophe de Sansal ◽

Benoit Devincre ◽

Ladislas P. Kubin

Keyword(s):

Plastic Deformation ◽

Mechanical Response ◽

Initial Density ◽

Three Dimensional ◽

Dislocation Dynamics ◽

Dynamics Simulation ◽

Key Factors ◽

Slip Mechanism ◽

Dynamics Simulations ◽

Dislocation Sources

This article reports on a study of the microstructure and mechanical response of copper polycrystals with grain sizes in the micrometer range. Three-dimensional dislocation dynamics simulations are used for the first time to investigate grain boundary strengthening and the Hall-Petch law. The methodology, which involves constructing a microcrystalline representative volume element with periodic boundary conditions, is briefly presented. Simulation results show that the initial density of dislocation sources and the cross-slip mechanism are two key factors controlling the heterogeneity of plastic deformation within the grains. At yield, the smaller the grains size, the more plastic deformation is heterogeneously distributed between grains and homogeneously distributed inside the grains. A size effect is reproduced and it is shown that the Hall-Petch exponent decreases from the very beginning of plastic flow and may reach a stable value at strains larger than the conventional proof stress.

Download Full-text

Practical electron tomography

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100140087 ◽

1995 ◽

Vol 53 ◽

pp. 742-743

Author(s):

C.L. Woodcock

Keyword(s):

Electron Tomography ◽

Tomographic Reconstruction ◽

Three Dimensional ◽

3D Shape ◽

Computational Resources ◽

Shape Of Particles

Despite the potential of the technique, electron tomography has yet to be widely used by biologists. This is in part related to the rather daunting list of equipment and expertise that are required. Thanks to continuing advances in theory and instrumentation, tomography is now more feasible for the non-specialist. One barrier that has essentially disappeared is the expense of computational resources. In view of this progress, it is time to give more attention to practical issues that need to be considered when embarking on a tomographic project. The following recommendations and comments are derived from experience gained during two long-term collaborative projects.Tomographic reconstruction results in a three dimensional description of an individual EM specimen, most commonly a section, and is therefore applicable to problems in which ultrastructural details within the thickness of the specimen are obscured in single micrographs. Information that can be recovered using tomography includes the 3D shape of particles, and the arrangement and dispostion of overlapping fibrous and membranous structures.

Download Full-text

Construction of plastic contact deformation maps on ceramics: a case study on aluminum nitride

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100165641 ◽

1996 ◽

Vol 54 ◽

pp. 636-637

Author(s):

D. L. Callahan

Keyword(s):

Plastic Deformation ◽

Aluminum Nitride ◽

Mechanical Response ◽

Three Dimensional ◽

Bright Field Image ◽

Perfectly Plastic ◽

Contrast Analysis ◽

Deformation Patterns

Modern polishing, precision machining and microindentation techniques allow the processing and mechanical characterization of ceramics at nanometric scales and within entirely plastic deformation regimes. The mechanical response of most ceramics to such highly constrained contact is not predictable from macroscopic properties and the microstructural deformation patterns have proven difficult to characterize by the application of any individual technique. In this study, TEM techniques of contrast analysis and CBED are combined with stereographic analysis to construct a three-dimensional microstructure deformation map of the surface of a perfectly plastic microindentation on macroscopically brittle aluminum nitride.The bright field image in Figure 1 shows a lg Vickers microindentation contained within a single AlN grain far from any boundaries. High densities of dislocations are evident, particularly near facet edges but are not individually resolvable. The prominent bend contours also indicate the severity of plastic deformation. Figure 2 is a selected area diffraction pattern covering the entire indentation area.

Download Full-text

Three-Dimensional reconstruction of isolated centrosomes by automated electron tomography

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100169286 ◽

1994 ◽

Vol 52 ◽

pp. 310-311

Author(s):

M.B. Braunfeld ◽

M. Moritz ◽

B.M. Alberts ◽

J.W. Sedat ◽

D.A. Agard

Keyword(s):

Electron Tomography ◽

Three Dimensional ◽

Vital Role ◽

Complex Nature ◽

Animal Cells ◽

Microtubule Organizing Center ◽

Nucleation Sites ◽

Dimensional Reconstruction ◽

Organizing Center ◽

Resolution Model

In animal cells, the centrosome functions as the primary microtubule organizing center (MTOC). As such the centrosome plays a vital role in determining a cell's shape, migration, and perhaps most importantly, its division. Despite the obvious importance of this organelle little is known about centrosomal regulation, duplication, or how it nucleates microtubules. Furthermore, no high resolution model for centrosomal structure exists.We have used automated electron tomography, and reconstruction techniques in an attempt to better understand the complex nature of the centrosome. Additionally we hope to identify nucleation sites for microtubule growth.Centrosomes were isolated from early Drosophila embryos. Briefly, after large organelles and debris from homogenized embryos were pelleted, the resulting supernatant was separated on a sucrose velocity gradient. Fractions were collected and assayed for centrosome-mediated microtubule -nucleating activity by incubating with fluorescently-labeled tubulin subunits. The resulting microtubule asters were then spun onto coverslips and viewed by fluorescence microscopy.

Download Full-text

Mechanical Behavior of Hydroxyapatite-Chitosan Composite: Effect of Processing Parameters

Minerals ◽

10.3390/min11020213 ◽

2021 ◽

Vol 11 (2) ◽

pp. 213

Author(s):

Hamid Ait Said ◽

Hassan Noukrati ◽

Hicham Ben Youcef ◽

Ayoub Bayoussef ◽

Hassane Oudadesse ◽

...

Keyword(s):

Molecular Weight ◽

Compressive Strength ◽

Mechanical Strength ◽

Bone Repair ◽

Mechanical Stability ◽

Three Dimensional ◽

Processing Parameters ◽

Composite Effect ◽

Solid Liquid ◽

The Impact

Three-dimensional hydroxyapatite-chitosan (HA-CS) composites were formulated via solid-liquid technic and freeze-drying. The prepared composites had an apatitic nature, which was demonstrated by X-ray diffraction and Infrared spectroscopy analyses. The impact of the solid/liquid (S/L) ratio and the content and the molecular weight of the polymer on the composite mechanical strength was investigated. An increase in the S/L ratio from 0.5 to 1 resulted in an increase in the compressive strength for HA-CSL (CS low molecular weight: CSL) from 0.08 ± 0.02 to 1.95 ± 0.39 MPa and from 0.3 ± 0.06 to 2.40 ± 0.51 MPa for the HA-CSM (CS medium molecular weight: CSM). Moreover, the increase in the amount (1 to 5 wt%) and the molecular weight of the polymer increased the mechanical strength of the composite. The highest compressive strength value (up to 2.40 ± 0.51 MPa) was obtained for HA-CSM (5 wt% of CS) formulated at an S/L of 1. The dissolution tests of the HA-CS composites confirmed their cohesion and mechanical stability in an aqueous solution. Both polymer and apatite are assumed to work together, giving the synergism needed to make effective cylindrical composites, and could serve as a promising candidate for bone repair in the orthopedic field.

Download Full-text